Records 1 - 50 / 384
Functional Divergence of Two Secreted Immune Proteases of Tomato
Ilyas, M. ; Hörger, A.C. ; Bozkurt, T.O. ; Burg, H.A. van den; Kaschani, F. ; Kaiser, M. ; Belhaj, K. ; Smoker, M. ; Joosten, M. ; Kamoun, S. ; Hoorn, R.A.L. van der - \ 2015
Current Biology 25 (2015)17. - ISSN 0960-9822 - p. 2300 - 2306.
cf-2-dependent disease resistance - pathogen effectors - transcription factors - provides insights - genome sequence - plant-pathogens - gene - defense - target - specialization
Rcr3 and Pip1 are paralogous secreted papain-like proteases of tomato. Both proteases are inhibited by Avr2 from the fungal pathogen Cladosporium fulvum, but only Rcr3 acts as a co-receptor for Avr2 recognition by the tomato Cf-2 immune receptor [ 1, 2, 3 and 4]. Here, we show that Pip1-depleted tomato plants are hyper-susceptible to fungal, bacterial, and oomycete plant pathogens, demonstrating that Pip1 is an important broad-range immune protease. By contrast, in the absence of Cf-2, Rcr3 depletion does not affect fungal and bacterial infection levels but causes increased susceptibility only to the oomycete pathogen Phytophthora infestans. Rcr3 and Pip1 reside on a genetic locus that evolved over 36 million years ago. These proteins differ in surface-exposed residues outside the substrate-binding groove, and Pip1 is 5- to 10-fold more abundant than Rcr3. We propose a model in which Rcr3 and Pip1 diverged functionally upon gene duplication, possibly driven by an arms race with pathogen-derived inhibitors or by coevolution with the Cf-2 immune receptor detecting inhibitors of Rcr3, but not of Pip1.
Novel introner-like elements in fungi are involved in parallel gains of spliceosomal introns
Collemare, J. ; Beenen, H.G. ; Crous, P.W. ; Wit, P.J.G.M. de; Burgt, A. van der - \ 2015
PLoS ONE 10 (2015)6. - ISSN 1932-6203 - 12 p.
daphnia populations - maximum-likelihood - evolution - gene - positions - conservation - selection - sequence - genomes
Spliceosomal introns are key components of the eukaryotic gene structure. Although they contributed to the emergence of eukaryotes, their origin remains elusive. In fungi, they might originate from the multiplication of invasive introns named Introner-Like Elements (ILEs). However, so far ILEs have been observed in six fungal species only, including Fulvia fulva and Dothistroma septosporum (Dothideomycetes), arguing against ILE insertion as a general mechanism for intron gain. Here, we identified novel ILEs in eight additional fungal species that are phylogenetically related to F. fulva and D. septosporum using PCR amplification with primers derived from previously identified ILEs. The ILE content appeared unique to each species, suggesting independent multiplication events. Interestingly, we identified four genes each containing two gained ILEs. By analysing intron positions in orthologues of these four genes in Ascomycota, we found that three ILEs had inserted within a 15 bp window that contains regular spliceosomal introns in other fungal species. These three positions are not the result of intron sliding because ILEs are newly gained introns. Furthermore, the alternative hypothesis of an inferred ancestral gain followed by independent losses contradicts the observed degeneration of ILEs. These observations clearly indicate three parallel intron gains in four genes that were randomly identified. Our findings suggest that parallel intron gain is a phenomenon that has been highly underestimated in ILE-containing fungi, and likely in the whole fungal kingdom.
Robustness to chronic heat stress in laying hens: a meta-analysis
Mignon-Grasteau, S. ; Moreri, U. ; Narcy, A. ; Rousseau, X. ; Rodenburg, T.B. ; Tixier-Boichard, M. ; Zerjal, T. - \ 2015
Poultry Science 94 (2015)4. - ISSN 0032-5791 - p. 586 - 600.
high ambient-temperatures - egg quality - naked neck - vitamin-c - performance - dwarf - gene - supplementation - management - nutrition
Chronic heat is a major stress factor in laying hens and many studies on the effect of heat stress have been published. It remains difficult, however, to draw general conclusions about the effect of chronic heat stress on performance and its relationship with genetic and environmental factors, as these studies have been done under varying experimental conditions and using various experimental designs. A meta-analysis enabled us to make a quantitative review of the results from 131 published papers. The relative effects of four factors (genotype, age, group size, and amplitude of temperature variation) and their interactions with temperature were analyzed for 13 traits. After pre-correcting the data for a random study effect, the best model for each trait was selected in a stepwise procedure based on its residual sum of squares. Shell strength, daily feed intake, egg mass, and hen-day egg production were found to be more sensitive to heat stress than the other traits as they dropped by 9.0 to 22.6% between thermo-neutrality (15 to 20°C) and heat stress (30 to 35°C) while yolk and albumen proportions or Haugh units showed nearly no variation with temperature (
Control of oriented cell division in the Arabidopsis embryo
Dop, M. van; Liao, C.Y. ; Weijers, D. - \ 2015
Current Opinion in Plant Biology 23 (2015). - ISSN 1369-5266 - p. 25 - 30.
preprophase band organization - plant development - transcription factor - monopteros - gene - root - differentiation - morphogenesis - cytokinesis - proteins
Multicellular plant development requires strict control of cell division orientation. A key unanswered question is how developmental regulators interact with the generic cell division machinery to trigger oriented divisions. We discuss the Arabidopsis embryo as a model for addressing this question. Recent progress in 3D imaging and computation now allows sketching of a framework for the developmental control of division orientation in which the signaling molecule auxin controls oriented division by preventing a geometrically defined default plane. We expect that the identification of auxin effectors, together with the identification of novel regulators of cell division will help to link developmental regulators to the division machinery.
Tumour necrosis factor allele variants and their association with the occurrence and severity of malaria in African children: a longitudinal study
Gichohi-Wainaina, W.N. ; Boonstra, A. ; Feskens, E.J.M. ; Demir, A.Y. ; Veenemans, J. ; Verhoef, H. - \ 2015
Malaria Journal 14 (2015). - ISSN 1475-2875 - 11 p.
plasmodium-falciparum malaria - tnf-alpha promoter - cerebral malaria - linkage disequilibrium - rheumatoid-arthritis - diabetes-mellitus - polymorphisms - gene - disease - hla
Background Tumour necrosis factor (TNF) is central to the immune response to Plasmodium infection. Its plasma concentration is influenced by allele variants in the promoter region of TNF. The study’s objectives were to assess TNF allele variants (TNF-1031 , TNF-308 ): (1) modulation of malaria rates in young Tanzanian children; (2) modulation of the severity of malaria as indicated by haemoglobin concentrations at the time of presentation with febrile episodes; and (3) the association between Plasmodium infection and haemoglobin concentration in symptomless parasite carriers. Methods Data from a placebo-controlled trial in which 612 Tanzanian children aged 6–60 months with height-for-age z-score in the range -3 SD to 1.5 SD was utilised. Those with Plasmodium infection at baseline were treated with artemether-lumefantrine. An episode of malaria was predefined as current Plasmodium infection with an inflammatory response (axillary temperature =37.5°C or whole blood C-reactive protein concentration =8 mg/L) in children reported sick. Linkage disequilibrium (LD) pattern assessment as well as haplotype analysis was conducted using HAPLOVIEW. Cox regression models used in the primary analysis accounted for multiple episodes per child. Results Genotyping of 94.9% (581/612) children for TNF-1031 (TNF-1031 T>C); allele frequency was 0.39. Corresponding values for rs1800629 (TNF-308 G>A) were 95.4% (584/612) and 0.17. Compared to the wild type genotype (TT), malaria rates were increased in the TNF-1031 CC genotype (hazard ratio, HR [95% CI]: 1.41 [1.01¿1.97] and 1.31 [0.97¿1.76] for crude analysis and adjusting for pre-specified baseline factors, respectively) but decreased in those with the TNF-308 AA genotype (corresponding HR: 0.13 [0.02¿0.63] and 0.16 [0.04¿0.67]). These associations were weaker when analysing first episodes of malaria (P value -0.59 and 0.38, respectively). No evidence that allele variants of TNF-1031 and TNF-308 affected haemoglobin concentration at first episode of malaria, or that they modified the association between Plasmodium infection and haemoglobin concentrations at baseline was observed.
Immune activation mediated by the late blight resistance protein R1 requires nuclear localization of R1 and AVR1
Du, Y. ; Berg, J. ; Govers, F. ; Bouwmeester, K. - \ 2015
New Phytologist 207 (2015)3. - ISSN 0028-646X - p. 735 - 747.
disease-resistance - phytophthora-infestans - arabidopsis-thaliana - innate immunity - plant immunity - receptor - recognition - potato - gene - component
Resistance against oomycete pathogens is mainly governed by intracellular nucleotide-binding leucine-rich repeat (NLR) receptors that recognize matching avirulence (AVR) proteins from the pathogen, RXLR effectors that are delivered inside host cells. Detailed molecular understanding of how and where NLR proteins and RXLR effectors interact is essential to inform the deployment of durable resistance (R) genes. Fluorescent tags, nuclear localization signals (NLSs) and nuclear export signals (NESs) were exploited to determine the subcellular localization of the potato late blight protein R1 and the Phytophthora infestans RXLR effector AVR1, and to target these proteins to the nucleus or cytoplasm. Microscopic imaging revealed that both R1 and AVR1 occurred in the nucleus and cytoplasm, and were in close proximity. Transient expression of NLS- or NES-tagged R1 and AVR1 in Nicotiana benthamiana showed that activation of the R1-mediated hypersensitive response and resistance required localization of the R1/AVR1 pair in the nucleus. However, AVR1-mediated suppression of cell death in the absence of R1 was dependent on localization of AVR1 in the cytoplasm. Balanced nucleocytoplasmic partitioning of AVR1 seems to be a prerequisite. Our results show that R1-mediated immunity is activated inside the nucleus with AVR1 in close proximity and suggest that nucleocytoplasmic transport of R1 and AVR1 is tightly regulated.
Remarkably divergent regions punctuate the genome assembly of the Caenorhabditis elegans Hawaiian strain CB4856
Thompson, O.A. ; Snoek, L.B. ; Nijveen, H. ; Sterken, M.G. ; Volkers, R.J.M. ; Brenchley, R. ; Hof, A. van 't; Bevers, R.P.J. ; Cossins, A.R. ; Yanai, I. ; Hajnal, A. ; Schmid, T. ; Perkins, J.D. ; Spencer, D. ; Kruglyak, L. ; Andersen, E.C. ; Moerman, D.G. ; Hillier, L.W. ; Kammenga, J.E. ; Waterston, R.H. - \ 2015
Genetics 200 (2015)3. - ISSN 0016-6731 - p. 975 - 989.
natural variation data - c. elegans - arabidopsis-thaliana - gene - polymorphism - populations - diversity - nematodes - dna - evolutionary
The Hawaiian strain (CB4856) of Caenorhabditis elegans is one of the most divergent from the canonical laboratory strain N2 and has been widely used in developmental, population and evolutionary studies. To enhance the utility of the strain, we have generated a draft sequence of the CB4856 genome, exploiting a variety of resources and strategies. The CB4856 genome when compared against the N2 reference has 327,050 single nucleotide variants (SNVs) and 79,529 insertion-deletion events (indels) that result in a total of 3.3 megabasepairs (Mb) of N2 sequence missing from CB4856 and 1.4 Mb of sequence present in CB4856 not present in N2. As previously reported, the density of SNVs varies along the chromosomes, with the arms of chromosomes showing greater average variation than the centers. In addition, we find 61 regions totaling 2.8 Mb, distributed across all six chromosomes, that have a greatly elevated SNV density, ranging from 2% to 16% SNVs. A survey of other wild isolates show that the two alternative haplotypes for each region are widely distributed, suggesting they have been maintained by balancing selection over long evolutionary times. These divergent regions contain an abundance of genes from large rapidly evolving families encoding F-box, MATH, BATH, seven-transmembrane G-coupled receptors, and nuclear hormone receptors suggesting that they provide selective advantages in natural environments. The draft sequence makes available a comprehensive catalog of sequence differences between the CB4856 and N2 strains that will facilitate the molecular dissection of their phenotypic differences. Our work also emphasizes the importance of going beyond simple alignment of reads to a reference genome when assessing differences between genomes.
Systemic Regulation of RAS/MAPK Signaling by the Serotonin Metabolite 5-HIAA
Schmid, T. ; Snoek, L.B. ; Fröhli, E. ; Bent, M.L. van der; Kammenga, J.E. ; Hajnal, A. - \ 2015
Plos Genetics 11 (2015)5. - ISSN 1553-7404 - 16 p.
caenorhabditis-elegans - c-elegans - natural variation - vulvar induction - complex disease - receptor - protein - gene - kinase - activation
Human cancer is caused by the interplay of mutations in oncogenes and tumor suppressor genes and inherited variations in cancer susceptibility genes. While many of the tumor initiating mutations are well characterized, the effect of genetic background variation on disease onset and progression is less understood. We have used C. elegans genetics to identify genetic modifiers of the oncogenic RAS/MAPK signaling pathway. Quantitative trait locus analysis of two highly diverged C. elegans isolates combined with allele swapping experiments identified the polymorphic monoamine oxidase A (MAOA) gene amx-2 as a negative regulator of RAS/MAPK signaling. We further show that the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA), which is a product of MAOA catalysis, systemically inhibits RAS/MAPK signaling in different organs of C. elegans. Thus, MAOA activity sets a global threshold for MAPK activation by controlling 5-HIAA levels. To our knowledge, 5-HIAA is the first endogenous small molecule that acts as a systemic inhibitor of RAS/MAPK signaling.
Effect of the DGAT1 K232A genotype of dairy cows on the milk metabolome and proteome
Lu, J. ; Boeren, S. ; Hooijdonk, A.C.M. van; Vervoort, J.J.M. ; Hettinga, K.A. - \ 2015
Journal of Dairy Science 98 (2015)5. - ISSN 0022-0302 - p. 3460 - 3469.
h-1-nmr spectroscopy - sample preparation - identification - stomatin - membrane - proteins - gene - cattle - yield
Diglyceride O-acyltransferase 1 (DGAT1) is the enzyme that catalyzes the synthesis of triglycerides from diglycerides and acyl-coenzyme A. The DGAT1 K232A polymorphism was previously shown to have a significant influence on bovine milk production characteristics (milk yield, protein content, fat content, and fatty acid composition). The mechanism of this influence has, however, not been elucidated. In this study, metabolomics (1H-nuclear magnetic resonance) and proteomics (laser chromatography-tandem mass spectrometry) were applied to determine the serum and lipid metabolite composition and milk fat globule membrane proteome of milk samples from cows with the DGAT1 KK and AA genotypes. The milk samples from cows with the DGAT1 KK genotype contained more stomatin, sphingomyelin, choline, and carnitine, and less citrate, creatine or phosphocreatine, glycerol-phosphocholine, mannose-like sugar, acetyl sugar phosphate, uridine diphosphate (UDP)-related sugar, and orotic acid compared with milk samples from cows with the DGAT1 AA genotype. Based on these results, we propose that the differences between the DGAT1 genotypes may be related to stomatin-sphingomyelin lipid rafts as well as structural (cell membrane) differences in epithelial cells of the mammary gland. In conclusion, our study shows that, in addition to previously described changes in triglyceride composition, cows differing in DGAT1 polymorphism differ in their milk proteome and metabolome, which may help in further understanding the effect of the DGAT1 K232A polymorphism on milk production characteristics.
Molecular cloning and characterization of the trichome specificchrysanthemyl diphosphate/chrysanthemol synthase promoter fromTanacetum cinerariifolium
Sultana, S. ; Hu, H. ; Gao, L. ; Mao, J. ; Luo, J. ; Jongsma, M.A. ; Wang, C. - \ 2015
Scientia Horticulturae 185 (2015). - ISSN 0304-4238 - p. 193 - 199.
mosaic virus-35s promoter - artemisia-annua - transcription factor - diphosphate synthase - pyrethrin biosynthesis - transgene expression - plant transformation - glandular trichomes - gene - cinerariaefolium
Natural pyrethrins accumulate to high concentrations in the flower achenes of pyrethrum (Tanacetumcinerariifolium). They are extracted from dried flowers and widely used as a natural pesticide. Chrysanthe-mol synthase (CHS) is the first key enzyme in the biosynthesis of pyrethrins. In this work, a 1128 bp TcCHSpromoter fragment was cloned from pyrethrum genomic DNA. The sequence contained cis-elementspredicted to be responsive to different hormones, light, and environmental stresses. To characterizethe promoter it was fused to the reporter genes green fluorescent protein (GFP) and -glucuronidase(GUS), and respectively transformed into florist’s chrysanthemum (Chrysanthemum morifolium ‘1581’)and tobacco (Nicotiana tabacum). GFP fluorescence in florist’s chrysanthemum ‘1581’and GUS staining oftobacco showed that the TcCHS promoter was exclusively expressed in the glandular secretory trichomes(GSTs) of both plant species. The findings will support research on factors influencing the accumulationof pyrethrins, and can be used for trichome-specific metabolic engineering of plants to ensure minimaladverse effects on plant growth and development.
The cell size distribution of tomato fruit can be changed by overexpression of CDKA1
Czerednik, A. ; Busscher, M. ; Angenent, G.C. ; Maagd, R.A. de - \ 2015
Plant Biotechnology Journal 13 (2015)2. - ISSN 1467-7644 - p. 259 - 268.
cyclin-dependent kinase - lycopersicon-esculentum mill - plant development - arabidopsis - endoreduplication - growth - gene - expression - division - dna
Tomato is one of the most cultivated vegetables in the world and an important ingredient of the human diet. Tomato breeders and growers face a continuous challenge of combining high quantity (production volume) with high quality (appearance, taste and perception for the consumers, processing quality for the processing industry). To improve the quality of tomato, it is important to understand the regulation of fruit development and of fruit cellular structure, which is in part determined by the sizes and numbers of cells within a tissue. The role of the cell cycle therein is poorly understood. Plant cyclin-dependent kinases (CDKs) are homologues of yeast cdc2, an important cell cycle regulator conserved throughout all eukaryotes. CDKA1 is constitutively expressed during the cell cycle and has dual functions in S- and M-phase progression. We have produced transgenic tomato plants with increased expression of CDKA1 under the control of the fruit-specific TPRP promoter, which despite a reduced number of seeds and diminished amount of jelly, developed fruits with weight and shape comparable to that of wild-type fruits. However, the phenotypic changes with regard to the pericarp thickness and placenta area were remarkable. Fruits of tomato plants with the highest expression of CDKA1 had larger septa and columella (placenta), compared with wild-type fruits. Our data demonstrate the possibility of manipulating the ratio between cell division and expansion by changing the expression of a key cell cycle regulator and probably its activity with substantial effects on structural traits of the harvested fruit.
Natural loss-of-function mutation of EDR1 conferring resistance to tomato powdery mildew in Arabidopsis thaliana accession C24
Gao, D. ; Appiano, M. ; Huibers, R.P. ; Loonen, A.E.H.M. ; Visser, R.G.F. ; Wolters, A.M.A. ; Bai, Y. - \ 2015
Molecular Plant Pathology 16 (2015)1. - ISSN 1464-6722 - p. 71 - 82.
salicylic-acid - downy mildew - gene - defense - plants - microsatellites - mechanism - evolution - cloning - kinase
To screen for potentially novel types of resistance to tomato powdery mildew Oidium neolycopersici, a disease assay was performed on 123 Arabidopsis thaliana accessions. Forty accessions were fully resistant, and one, C24, was analysed in detail. By quantitative trait locus (QTL) analysis of an F2 population derived from C24 × Sha (susceptible accession), two QTLs associated with resistance were identified in C24. Fine mapping of QTL-1 on chromosome 1 delimited the region to an interval of 58¿kb encompassing 15 candidate genes. One of these was Enhanced Disease Resistance 1 (EDR1). Evaluation of the previously obtained edr1 mutant of Arabidopsis accession Col-0, which was identified because of its resistance to powdery mildew Golovinomyces cichoracearum, showed that it also displayed resistance to O.¿neolycopersici. Sequencing of EDR1 in our C24 germplasm (referred to as C24-W) revealed two missing nucleotides in the second exon of EDR1 resulting in a premature stop codon. Remarkably, C24 obtained from other laboratories does not contain the EDR1 mutation. To verify the identity of C24-W, a DNA region containing a single nucleotide polymorphism (SNP) unique to C24 was sequenced showing that C24-W contains the C24-specific nucleotide. C24-W showed enhanced resistance to O.¿neolycopersici compared with C24 not containing the edr1 mutation. Furthermore, C24-W displayed a dwarf phenotype, which was not associated with the mutation in EDR1 and was not caused by the differential accumulation of pathogenesis-related genes. In conclusion, we identified a natural edr1 mutant in the background of C24.
Characterization of apoptosis in PER.C6® batch and perfusion cultures
Mercier, S.M. ; Diepenbroek, B. ; Martens, D.E. ; Wijffels, R.H. ; Streefland, M. - \ 2015
Biotechnology and Bioengineering 112 (2015)3. - ISSN 0006-3592 - p. 569 - 578.
hamster ovary cells - high-level expression - flow-cytometry - line per.c6 - death - adenovirus - dna - vaccine - gene - manufacture
Preventing or delaying cell death is a challenge in mammalian cell cultures for the development and optimization of production processes for biopharmaceuticals. Cell cultures need to be maintained highly viable for extended times in order to reach maximum production yields. Moreover, programmed cell death through apoptosis is often believed to occur without being detected by classical viability measurements. In this study, we characterized cell death in PER.C6® batch and perfusion cultures using three flow cytometry techniques measuring different steps of the apoptosis cascade: DNA fragmentation, caspases activation and phosphatidylserine externalization. We showed that apoptosis is the main pathway of PER.C6® cell death in batch cultures after depletion of main carbon sources. In high cell density perfusion cultures fed at a constant specific perfusion rate, both high viability and very limited apoptosis were observed. When extending this perfusion process far beyond standard operations, cultures were exposed to suboptimal process conditions, which resulted in an increase of apoptotic cell death. Moreover, we showed that the reference viability measurement using trypan blue exclusion properly assesses the level of cell death in PER.C6® cultures. This study is a first step in understanding the mechanisms of PER.C6® cell death, which will be helpful to support applications of the cell line.
Population structure and pathotype diversity of the wheat blast pathogen Magnaporthe oryzae 25 years after its emergence in Brazil
Nunes Maciel, J.L. ; Ceresini, P. ; Castroagudin, V.L. ; Zala, M. ; Kema, G.H.J. - \ 2014
Phytopathology 104 (2014)1. - ISSN 0031-949X - p. 95 - 107.
mating-type distribution - pyricularia-grisea - mycosphaerella-graminicola - genotypic diversity - fertility status - growth-stages - rice - resistance - gene - recombination
Since its first report in Brazil in 1985, wheat blast, caused by Magnaporthe oryzae (anamorph: Pyricularia oryzae), has become increasingly important in South America, where the disease is still spreading. We used 11 microsatellite loci to elucidate the population structure of the wheat blast pathogen in wheat fields in central-western, southeastern, and southern Brazil. No subdivision was found among the wheat-infecting populations, consistent with high levels of gene flow across a large spatial scale. Although the clonal fraction was relatively high and the two mating type idiomorphs (MAT1-1 and MAT1-2) were not at similar frequencies, the clone-corrected populations from Distrito Federal and Goiás, Minas Triangle, and São Paulo were in gametic equilibrium. Based on these findings, we propose that populations of the wheat blast pathogen exhibit a mixed reproductive system in which sexual reproduction is followed by the local dispersal of clones. Seedling virulence assays with local wheat cultivars differentiated 14 pathotypes in the current population. Detached head virulence assays differentiated eight virulence groups on the same wheat cultivars. There was no correlation between seedling and head reaction
KORRIGAN1 Interacts Specifically with Integral Components of the Cellulose Synthase Machinery
Mansoori Zangir, N. ; Timmers, J.F.P. ; Desprez, T. ; Lessa Alvim Kamei, C. ; Dees, D.C.T. ; Vincken, J.P. ; Visser, R.G.F. ; Höfte, H. ; Vernhettes, S. ; Trindade, L.M. - \ 2014
PLoS ONE 9 (2014)11. - ISSN 1932-6203
secondary cell-wall - arabidopsis-thaliana - endo-1,4-beta-glucanase - expression - membranes - protein - system - plants - gene - endo-1,4-beta-d-glucanase
Cellulose is synthesized by the so called rosette protein complex and the catalytic subunits of this complex are the cellulose synthases (CESAs). It is thought that the rosette complexes in the primary and secondary cell walls each contains at least three different non-redundant cellulose synthases. In addition to the CESA proteins, cellulose biosynthesis almost certainly requires the action of other proteins, although few have been identified and little is known about the biochemical role of those that have been identified. One of these proteins is KORRIGAN (KOR1). Mutant analysis of this protein in Arabidopsis thaliana showed altered cellulose content in both the primary and secondary cell wall. KOR1 is thought to be required for cellulose synthesis acting as a cellulase at the plasma membrane–cell wall interface. KOR1 has recently been shown to interact with the primary cellulose synthase rosette complex however direct interaction with that of the secondary cell wall has never been demonstrated. Using various methods, both in vitro and in planta, it was shown that KOR1 interacts specifically with only two of the secondary CESA proteins. The KOR1 protein domain(s) involved in the interaction with the CESA proteins were also identified by analyzing the interaction of truncated forms of KOR1 with CESA proteins. The KOR1 transmembrane domain has shown to be required for the interaction between KOR1 and the different CESAs, as well as for higher oligomer formation of KOR1.
Accuracy of genomic prediction when combining two related crossbred populations
Vallee, A.A.A. ; Arendonk, J.A.M. van; Bovenhuis, H. - \ 2014
Journal of Animal Science 92 (2014)10. - ISSN 0021-8812 - p. 4342 - 4348.
dairy-cattle breeds - beef-cattle - selection - performance - animals - values - uterine - traits - impact - gene
Charolais bulls are selected for their crossbreed performance when mated to Montbéliard or Holstein dams. To implement genomic prediction, one could build a reference population for each crossbred population independently. An alternative could be to combine both crossbred populations into a single reference population to increase size and accuracy of prediction. The objective of this study was to investigate the accuracy of genomic prediction by combining different crossbred populations. Three scenarios were considered: 1) using 1 crossbred population as reference to predict phenotype of animals from the same crossbred population, 2) combining the 2 crossbred populations into 1 reference to predict phenotype of animals from 1 crossbred population, and 3) using 1 crossbred population as reference to predict phenotype of animals from the other crossbred population. Traits studied were bone thinness, height, and muscular development. Phenotypes and 45,117 SNP genotypes were available for 1,764 Montbéliard × Charolais calves and 447 Holstein × Charolais calves. The population was randomly spilt into 10 subgroups, which were assigned to the validation one by one. To allow fair comparison between scenarios, size of the reference population was kept constant for all scenarios. Breeding values were estimated with BLUP and genomic BLUP. Accuracy of prediction was calculated as the correlation between the EBV and the phenotypic values of the calves in the validation divided by the square root of the heritability. Genomic BLUP showed higher accuracies (between 0.281 and 0.473) than BLUP (between 0.197 and 0.452). Accuracies tended to be highest when prediction was within 1 crossbred population, intermediate when populations were combined into the reference population, and lowest when prediction was across populations. Decrease in accuracy from a prediction within 1 population to a prediction across populations was more pronounced for bone thinness (–27%) and height (–29%) than for muscular development (–14%). Genetic correlation between the 2 crossbred populations was estimated using pedigree relationships. It was 0.70 for bone thinness, 0.80 for height, and 0.99 for muscular development. Genetic correlation indicates the expected gain in accuracy of prediction when combining different populations into 1 reference population. The larger the genetic correlation is, the larger the benefit is to combine populations for genomic prediction.
Precise control of plant stem cell activity through parallel regulatory inputs
Bennett, T. ; Toorn, A. van; Willemsen, V. ; Scheres, B. - \ 2014
Development 141 (2014). - ISSN 0950-1991 - p. 4055 - 4064.
arabidopsis-thaliana root - transcription factor - shoot apex - meristem - gene - differentiation - organization - maintenance - homeostasis - sombrero
The regulation of columella stem cell activity in the Arabidopsis root cap by a nearby organizing centre, the quiescent centre, has been a key example of the stem cell niche paradigm in plants. Here, we investigate interactions between transcription factors that have been shown to regulate columella stem cells using a simple quantification method for stem cell activity in the root cap. Genetic and expression analyses reveal that the RETINOBLASTOMA-RELATED protein, the FEZ and SOMBRERO NAC-domain transcription factors, the ARF10 and ARF16 auxin response factors and the quiescent centre-expressed WOX5 homeodomain protein each provide independent inputs to regulate the number of columella stem cells. Given the tight control of columella development, we found that these inputs act in a surprisingly parallel manner. Nevertheless, important points of interaction exist; for example, we demonstrate the repression of SMB activity by non-autonomous action of WOX5. Our results suggest that the developmental progression of columella stem cells may be quantitatively regulated by several more broadly acting transcription factors rather than by a single intrinsic stem cell factor, which raises questions about the special nature of the stem cell state in plants.
Toolkit for Visualization of the Cellular Structure and Organelles in Aspergillus niger
Buren, E.B.J. ten; Karrenbelt, M.A.P. ; Lingemann, M. ; Chordia, S. ; Deng, Y. ; Hu, J.J. ; Verest, J.M. ; Wu, V. ; Bello Gonzalez, T.D.G. ; Heck, R.G.A. van; Odoni, D.I. ; Schonewille, T. ; Straat, L. van der; Graaff, L.H. de; Passel, M.W.J. van - \ 2014
ACS synthetic biology 3 (2014)12. - ISSN 2161-5063 - p. 995 - 998.
gene - hyphae
Aspergillus niger is a filamentous fungus that is extensively used in industrial fermentations for protein expression and the production of organic acids. Inherent biosynthetic capabilities, such as the capacity to secrete these biomolecules in high amounts, make A. niger an attractive production host. Although A. niger is renowned for this ability, the knowledge of the molecular components that underlie its production capacity, intercellular trafficking processes and secretion mechanisms is far from complete. Here, we introduce a standardized set of tools, consisting of an N-terminal GFP-actin fusion and codon optimized eforRed chromoprotein. Expression of the GFP-actin construct facilitates visualization of the actin filaments of the cytoskeleton, whereas expression of the chromoprotein construct results in a clearly distinguishable red phenotype. These experimentally validated constructs constitute the first set of standardized A. niger biomarkers, which can be used to study morphology, intercellular trafficking, and secretion phenomena.
REDUCED DORMANCY5 Encodes a Protein Phosphatase 2C That Is Required for Seed Dormancy in Arabidopsis
Xiang, Y. ; Nakabayashi, K. ; Ding, J. ; He, F. ; Bentsink, L. ; Soppe, W.J.J. - \ 2014
The Plant Cell 26 (2014)11. - ISSN 1040-4651 - p. 4362 - 4375.
rna-binding proteins - abscisic-acid - messenger-rna - pp2c phosphatases - germination - thaliana - aba - reveals - gene - mutants
Seed dormancy determines germination timing and contributes to crop production and the adaptation of natural populations to their environment. Our knowledge about its regulation is limited. In a mutagenesis screen of a highly dormant Arabidopsis thaliana line, the reduced dormancy5 (rdo5) mutant was isolated based on its strongly reduced seed dormancy. Cloning of RDO5 showed that it encodes a PP2C phosphatase. Several PP2C phosphatases belonging to clade A are involved in abscisic acid signaling and control seed dormancy. However, RDO5 does not cluster with clade A phosphatases, and abscisic acid levels and sensitivity are unaltered in the rdo5 mutant. RDO5 transcript could only be detected in seeds and was most abundant in dry seeds. RDO5 was found in cells throughout the embryo and is located in the nucleus. A transcriptome analysis revealed that several genes belonging to the conserved PUF family of RNA binding proteins, in particular Arabidopsis PUMILIO9 (APUM9) and APUM11, showed strongly enhanced transcript levels in rdo5 during seed imbibition. Further transgenic analyses indicated that APUM9 reduces seed dormancy. Interestingly, reduction of APUM transcripts by RNA interference complemented the reduced dormancy phenotype of rdo5, indicating that RDO5 functions by suppressing APUM transcript levels.
Expression of natural human b1,4-GalT1 variants and of non-mammalian homologues in plants leads to differences in galactosylation of N-glycans
Hesselink, T. ; Rouwendal, G.J.A. ; Henquet, M.G.L. ; Florack, D.E.A. ; Helsper, J.P.F.G. ; Bosch, H.J. - \ 2014
Transgenic Research 23 (2014)5. - ISSN 0962-8819 - p. 717 - 728.
golgi-apparatus - murine beta-1,4-galactosyltransferase - beta 1,4-galactosyltransferase - transgenic plants - gene - cells - localization - antibodies - oligosaccharides - glycoproteins
b1,4-Galactosylation of plant N-glycans is a prerequisite for commercial production of certain biopharmaceuticals in plants. Two different types of galactosylated N-glycans have initially been reported in plants as the result of expression of human b1,4-galactosyltransferase 1 (GalT). Here we show that these differences are associated with differences at its N-terminus: the natural short variant of human GalT results in hybrid type N-glycans, whereas the long form generates bi-antennary complex type N-glycans. Furthermore, expression of non-mammalian, chicken and zebrafish GalT homologues with N-termini resembling the short human GalT N-terminus also induce hybrid type N-glycans. Providing both non-mammalian GalTs with a 13 amino acid N-terminal extension that distinguishes the two naturally occurring forms of human GalT, acted to increase the levels of biantennary galactosylated N-glycans when expressed in tobacco leaves. Replacement of the cytosolic tail and transmembrane domain of chicken and zebrafish GalTs with the corresponding region of rat a2,6-sialyltransferase yielded a gene whose expression enhanced the level of bi-antennary galactosylation even further.
BioHackathon series in 2011 and 2012: penetration of ontology and linked data in life science domains
Katayama, T. ; Wilkinson, M.D. ; Aoki-Kinoshita, K.F. ; Prins, J.C.P. - \ 2014
Journal of Biomedical Semantics 5 (2014). - ISSN 2041-1480
genome analysis environment - metabolic pathways - web services - gene - sequences - software - biology - normalization - collection - glycomics
The application of semantic technologies to the integration of biological data and the interoperability of bioinformatics analysis and visualization tools has been the common theme of a series of annual BioHackathons hosted in Japan for the past five years. Here we provide a review of the activities and outcomes from the BioHackathons held in 2011 in Kyoto and 2012 in Toyama. In order to efficiently implement semantic technologies in the life sciences, participants formed various sub-groups and worked on the following topics: Resource Description Framework (RDF) models for specific domains, text mining of the literature, ontology development, essential metadata for biological databases, platforms to enable efficient Semantic Web technology development and interoperability, and the development of applications for Semantic Web data. In this review, we briefly introduce the themes covered by these sub-groups. The observations made, conclusions drawn, and software development projects that emerged from these activities are discussed.
A weighted AMMI Algorithm to Study Genotype-by-Environment Interaction and QTL-by-Environment Interaction
Rodrigues, P.C. ; Malosetti, M. ; Gauch, H.G. - \ 2014
Crop Science 54 (2014)4. - ISSN 0011-183X - p. 1555 - 1570.
principal component analysis - multiplicative interaction-model - joint regression-analysis - additive main - cross-validation - yield trials - barley cross - mixed-model - selection - gene
Genotype-by-environment (G × E) interaction (GEI) and quantitative trait locus (QTL)-by-environment interaction (QEI) are common phenomena in multiple-environment trials and represent a major challenge to breeders. The additive main effects and multiplicative interaction (AMMI) model is a widely used tool for the analysis of multiple-environment trials, where the data are represented by a two-way table of G × E means. For complete tables, least squares estimation for the AMMI model is equivalent to fitting an additive two-way ANOVA model for the main effects and applying a singular value decomposition to the interaction residuals, thereby implicitly assuming equal weights for all G × E means. However, multiple-environment data with strong GEI are often also characterized by strong heterogeneous error variation. To improve the performance of the AMMI model in the latter situation, we introduce a generalized estimation scheme, the weighted AMMI or W-AMMI algorithm. This algorithm is useful for studying GEI and QEI. For QEI, the W-AMMI algorithm can be used to create predicted values per environment that are subjected to QTL analysis. We compare the performance of this combined W-AMMI and QTL mapping strategy to direct QTL mapping on G × E means and to QTL mapping on AMMI-predicted values, again with QTL analyses for individual environments. Finally, we compare the W-AMMI QTL mapping strategy, with a multi-environment mixed model QTL mapping approach. Two data sets are used: (i) data from a simulated pepper (Capsicum annuum L.) back cross population using a crop growth model to relate genotypes to phenotypes in a nonlinear way, and (ii) the doubled-haploid Steptoe × Morex barley (Hordeum vulgare L.) population. The QTL analyses on the W-AMMI-predicted values outperformed the QTL analyses on the G × E means and on the AMMI-predicted values, and were very similar to the mixed model QTL mapping approach with regard to the number and location of the true positive QTLs detected, especially for QTLs associated with the interaction and for environments with higher error variance. W-AMMI analysis for GEI and QEI provides an easy-to-use and robust tool with wide applicability.
Adipose tissue metabolism and inflammation are differently affected by weight loss in obese mice due to either a high-fat diet restriction or change to a low-fat diet
Hoevenaars, F.P.M. ; Keijer, J. ; Herreman, L. ; Palm, I.F. ; Hegeman, M.A. ; Swarts, J.J.M. ; Schothorst, E.M. van - \ 2014
Genes & Nutrition 9 (2014). - ISSN 1555-8932 - 11 p.
nicotinamide nucleotide transhydrogenase - c57bl/6j mice - mitochondrial biogenesis - energy restriction - insulin-resistance - glucose - acid - expression - health - gene
Restriction of a high-fat diet (HFD) and a change to a low-fat diet (LFD) are two interventions that were shown to promote weight loss and improve parameters of metabolic health in obesity. Examination of the biochemical and molecular responses of white adipose tissue (WAT) to these interventions has not been performed so far. Here, male C57BL/6JOlaHsd mice, harboring an intact nicotinamide nucleotide transhydrogenase gene, were fed a purified 40 energy% HFD for 14 weeks to induce obesity. Afterward, mice were divided into three dietary groups: HFD (maintained on HFD), LFD (changed to LFD with identical ingredients), and HFD-CR (restricted to 70 % of the HFD). The effects of the interventions were examined after 5 weeks. Beneficial effects were seen for both HFD-CR and LFD (compared to HFD) regarding physiological parameters (body weight and fat mass) and metabolic parameters, including circulating insulin and leptin levels. Macrophage infiltration in WAT was reduced by both interventions, although more effectively by HFDCR. Strikingly, molecular parameters in WAT differed between HFD-CR and LFD, with increased activation of mitochondrial carbohydrate and fat metabolism in HFDCR mice. Our results confirm that restriction of the amount of dietary intake and reduction in the dietary energy content are both effective in inducing weight loss. The larger decrease in WAT inflammation and increase in mitochondrial carbohydrate metabolism may be due to a larger degree of energy restriction in HFD-CR, but could also be due to superior effectiveness of dietary restriction in weight loss strategies.
Analysis of the A-U rich hairpin from the intergenic region of tospovirus S RNA as target and inducer of RNA silencing
Hedil, M. ; Hassani-Mehraban, A. ; Lohuis, D. ; Kormelink, R.J.M. - \ 2014
PLoS ONE 9 (2014)9. - ISSN 1932-6203 - 10 p.
spotted-wilt-virus - small interfering rnas - double-stranded-rna - viral suppressors - gene - protein - plants - sequence - arabidopsis - resistance
Earlier work indicated that Tomato spotted wilt virus (TSWV) messenger transcripts, and not the (anti)genomic RNAs, are targeted by the RNA silencing machinery. Here, the predicted AU-rich hairpin (HP) structure encoded by the intergenic region (IGR) of the TSWV S RNA, and present at the 3' end of viral mRNAs, was analyzed as a target and inducer for RNA silencing. Virus-derived siRNAs (vsiRNAs) purified from virus infected plants were found to derive from all three genomic RNA segments but predominantly the ambisense M and S RNAs. Further profiling on the S RNA sequence revealed that vsiRNAs were found from almost the entire S RNA sequence, except the IGR from where hardly any vsiRNAs were found. Similar profiles were observed with the distantly related Tomato yellow ring tospovirus (TYRV). Dicer cleavage assays using Drosophila melanogaster (Dm) embryo extracts showed that synthetic transcripts of the IGR-HP region were recognized as substrate for Dicer. Transient agroinfiltration assays of a GFP-sensor construct containing the IGR-HP sequence at its 3' UTR (GFP-HP) did not show more rapid/strong silencing and profiling of the corresponding siRNAs, generated outside the context of a viral infection, still revealed relatively low levels of IGR-HP-derived siRNAs. These data support the idea that the IGR-HP is a weak inducer of RNA silencing and only plays a minor role in the amplification of a strong antiviral RNAi response.
Phenotypic analyses of Arabidopsis T-DNA insertion lines and expression profiling reveal that multiple L-type lectin receptor kinases are involved in plant immunity
Wang, Y. ; Bouwmeester, K. ; Beseh, P. ; Shan, W. ; Govers, F. - \ 2014
Molecular Plant-Microbe Interactions 27 (2014)12. - ISSN 0894-0282 - p. 1390 - 1402.
pattern-triggered immunity - phytophthora-infestans - salicylic-acid - defense responses - innate immunity - thaliana - gene - resistance - biology - roles
L-type lectin receptor kinases (LecRKs) are membrane-spanning receptor-like kinases with putative roles in biotic and abiotic stress responses and in plant development. In Arabidopsis, 45 LecRKs were identified but their functions are largely unknown. Here, a systematic functional analysis was carried out by evaluating phenotypic changes of Arabidopsis LecRK T-DNA insertion lines in plant development and upon exposure to various external stimuli. None of the LecRK T-DNA insertion lines showed clear developmental changes, neither under normal conditions nor upon abiotic stress treatment. However, many of the T-DNA insertion lines showed altered resistance to Phytophthora brassicae, Phytophthora capsici, Pseudomonas syringae or Alternaria brassicicola. One mutant defective in LecRK-V.5 expression, was compromised in resistance to two Phytophthora spp. but showed enhanced resistance to P. syringae. LecRK-V.5 overexpression confirmed its dual role in resistance and susceptibility depending on the pathogen. Combined analysis of these phenotypic data and LecRK expression profiles retrieved from public datasets revealed that LecRKs which are hardly induced upon infection or even suppressed are also involved in pathogen resistance. Computed co-expression analysis revealed that LecRKs with similar function displayed diverse expression patterns. Since LecRKs are widespread in plants, the results presented here provide invaluable information for exploring the potential of LecRKs as novel sources of resistance in crops.
A strategy for developing representative germplasm sets for systematic QTL validation, demonstrated for apple, peach, and sweet cherry
Peace, C.P. ; Luby, J. ; Weg, W.E. van de; Bink, M.C.A.M. ; Iezzoni, A.F. - \ 2014
Tree Genetics and Genomes 10 (2014)6. - ISSN 1614-2942 - p. 1679 - 1694.
x domestica borkh. - marker-assisted selection - fire blight resistance - breeding program - population-structure - pyrus-communis - fruit firmness - map position - md-acs1 - gene
Horticultural crop improvement would benefit from a standardized, systematic, and statistically robust procedure for validating quantitative trait loci (QTLs) in germplasm relevant to breeding programs. Here, we describe and demonstrate a strategy for developing reference germplasm sets of perennial, clonally propagated crops, especially those with long juvenile periods. Germplasm is chosen to efficiently represent important members of larger pedigree-connected genepools. To facilitate validation of multiple QTLs, genome-wide representation of alleles is optimized for designated important breeding parents (IBPs) by estimating average allelic representation in relatives. The strategy and arising principles were demonstrated in a simulated germplasm set. Strong statistical power can be achieved with a carefully chosen germplasm set composed of IBPs, their numerous unselected progenies and close relatives, and all available founders and intermediate ancestors. Crop Reference Sets were developed in the marker-assisted breeding (MAB)-enabling “RosBREED” project as a base resource for QTL validation in US breeding germplasm of apple (Malus × domestica), peach (Prunus persica), and sweet cherry (Prunus avium) consisting of 467, 452, and 268 individuals, respectively. These sets adequately represent the most designated IBPs, have distinct advantages for QTL validation over other germplasm arrangements of equal size, and are recommended as a base resource for QTL validation by breeders of these US crops. The strategy described here can be used to develop efficient reference germplasm sets suiting other breeding genepools or to calculate the statistical power for QTL validation of germplasm sets already established.
Circulation of four Anaplasma phagocytophilum ecotypes in Europe
Jahfari, S. ; Coipan, E.C. ; Fonville, M. ; Leeuwen, A.D. van; Hengeveld, P. ; Heylen, D. ; Heyman, P. ; Maanen, C. van; Butler, C.M. ; Foldvari, G. ; Szekeres, S. ; Duijvendijk, L.A.G. van; Tack, W. ; Rijks, J.M. ; Giessen, J. van der; Takken, W. ; Wieren, S.E. van; Takumi, K. ; Sprong, H. - \ 2014
Parasites & Vectors 7 (2014)1. - ISSN 1756-3305
candidatus neoehrlichia mikurensis - human granulocytic anaplasmosis - ixodes-ricinus ticks - borrelia-burgdorferi - borne diseases - phylogenetic analyses - sequence-analysis - ehrlichiosis - strains - gene
Background: Anaplasma phagocytophilum is the etiological agent of granulocytic anaplasmosis in humans and animals. Wild animals and ticks play key roles in the enzootic cycles of the pathogen. Potential ecotypes of A. phagocytophilum have been characterized genetically, but their host range, zoonotic potential and transmission dynamics has only incompletely been resolved. Methods. The presence of A. phagocytophilum DNA was determined in more than 6000 ixodid ticks collected from the vegetation and wildlife, in 289 tissue samples from wild and domestic animals, and 69 keds collected from deer, originating from various geographic locations in The Netherlands and Belgium. From the qPCR-positive lysates, a fragment of the groEL-gene was amplified and sequenced. Additional groEL sequences from ticks and animals from Europe were obtained from GenBank, and sequences from human cases were obtained through literature searches. Statistical analyses were performed to identify A. phagocytophilum ecotypes, to assess their host range and their zoonotic potential. The population dynamics of A. phagocytophilum ecotypes was investigated using population genetic analyses. Results: DNA of A. phagocytophilum was present in all stages of questing and feeding Ixodes ricinus, feeding I. hexagonus, I. frontalis, I. trianguliceps, and deer keds, but was absent in questing I. arboricola and Dermacentor reticulatus. DNA of A. phagocytophilum was present in feeding ticks and tissues from many vertebrates, including roe deer, mouflon, red foxes, wild boar, sheep and hedgehogs but was rarely found in rodents and birds and was absent in badgers and lizards. Four geographically dispersed A. phagocytophilum ecotypes were identified, that had significantly different host ranges. All sequences from human cases belonged to only one of these ecotypes. Based on population genetic parameters, the potentially zoonotic ecotype showed significant expansion. Conclusion: Four ecotypes of A. phagocytophilum with differential enzootic cycles were identified. So far, all human cases clustered in only one of these ecotypes. The zoonotic ecotype has the broadest range of wildlife hosts. The expansion of the zoonotic A. phagocytophilum ecotype indicates a recent increase of the acarological risk of exposure of humans and animals.
Soybean SAT1 (Symbiotic Ammonium Transporter 1) encodes a bHLH transcription factor involved in nodule growth and NH4+ transport
Chiasson, D.M. ; Loughlin, P.C. ; Mazurkiewicz, D. ; Mohammadidehcheshmeh, M. ; Fedorova, E.E. ; Okamoto, M. ; McLean, E. ; Glass, A.D.M. ; Smith, S.E. ; Bisseling, T. ; Tyerman, S.D. ; Day, D.A. ; Kaiser, B.N. - \ 2014
Proceedings of the National Academy of Sciences of the United States of America 111 (2014)13. - ISSN 0027-8424 - p. 4814 - 4819.
arabidopsis-thaliana - circadian clock - lotus-japonicus - stress-response - er stress - membrane - protein - expression - domain - gene
Glycine max symbiotic ammonium transporter 1 was first documented as a putative ammonium (NH4+) channel localized to the symbiosome membrane of soybean root nodules. We show that Glycine max symbiotic ammonium transporter 1 is actually a membrane-localized basic helix–loop–helix (bHLH) DNA-binding transcription factor now renamed Glycine max bHLH membrane 1 (GmbHLHm1). In yeast, GmbHLHm1 enters the nucleus and transcriptionally activates a unique plasma membrane NH4+ channel Saccharomyces cerevisiae ammonium facilitator 1. Ammonium facilitator 1 homologs are present in soybean and other plant species, where they often share chromosomal microsynteny with bHLHm1 loci. GmbHLHm1 is important to the soybean rhizobium symbiosis because loss of activity results in a reduction of nodule fitness and growth. Transcriptional changes in nodules highlight downstream signaling pathways involving circadian clock regulation, nutrient transport, hormone signaling, and cell wall modification. Collectively, these results show that GmbHLHm1 influences nodule development and activity and is linked to a novel mechanism for NH4+ transport common to both yeast and plants.
Regeneration and transformation of Crambe abyssinica
Qi, W. ; Tinnenbroek-Capel, I.E.M. ; Schaart, J. ; Huang Bangquan, ; Cheng, J. ; Visser, R.G.F. ; Loo, E.N. van; Krens, F.A. - \ 2014
BMC Plant Biology 14 (2014). - ISSN 1471-2229 - 12 p.
gene - agrobacterium - tissue - plants
Background: Crambe abyssinica (crambe) is a non-food oil seed crop. Its seed oil is widely used in the chemical industry because of the high erucic acid content. Furthermore, it is a potential platform for various feedstock oils for industrial uses based on genetic modification. Here, we describe the development of a series of protocols for all steps required in the process of generating genetically modified crambe. Results: Different explant types from crambe seedlings were tested for shoot regeneration using different hormone-combinations. Cotyledonary nodes on basic medium with 0.5 µM NAA and 2.2 µM BAP gave the highest regeneration percentages. For propagation by tissue culture, explants of stems, petioles, leaves and axillary buds of in vitro plantlets were tested using the optimized medium. Axillary buds showed the highest shoot proliferation efficiency. Cotyledonary nodes were used to test the proper concentration of kanamycin for selection of transformation events, and 10 to 25 mg · L-1 were identified as effective. The cotyledonary nodes and cotyledons from 7-day-old seedlings were used in Agrobacterium-mediated transformations with two kinds of selection strategies, shifting or consistent. Using the shifting selection method (10 mg · L-1 kanamycin, 25 mg · L-1, then back to 10 mg · L-1) cotyledonary nodes gave 10% transformation frequency, and cotyledons 4%, while with the consistent method (25 mg · L-1) lower frequencies were found, 1% for cotyledonary nodes and 0% for cotyledons). Later, in vitro plant axillary buds were tried as explants for transformation, however, transformation frequency was low ranging from 0.5 to 2%. Overall, testing six different vectors and two kinds of Agrobacterium strains, the average transformation frequency using the shifting method was 4.4%. Determining T-DNA insertion numbers by Southern blotting showed that approximately 50% of the transgenic lines had a single-copy insertion. Conclusions: Present research revealed the potential of using crambe meristematic tissue for genetic transformation andin vitro propagation. The most efficient method of transformation used cotyledonary node explants from 7-days-old seedlings with a shifting kanamycin selection. Meristematic tissues (cotyledonary node or axillary bud) had the highest ability for shoot proliferation. Single-copy T-DNA insert lines could be efficiently and reproducibly generated.
Tomato yellow leaf curl virus resistance by ty-1 involves increased cytosine methylation of viral genomes and is compromised by cucumber mosaic virus infection
Butterbach, P.B.E. ; Verlaan, M.G. ; Dullemans, A.M. ; Lohuis, H. ; Visser, R.G.F. ; Bai, Y. ; Kormelink, R.J.M. - \ 2014
Proceedings of the National Academy of Sciences of the United States of America 111 (2014)35. - ISSN 0027-8424 - p. 12942 - 12947.
short interfering rna - dna methylation - geminivirus al2 - l2 proteins - adenosine kinase - gene - suppression - arabidopsis - plants - locus
Tomato yellow leaf curl virus (TYLCV) and related begomoviruses are a major threat to tomato production worldwide and, to protect against these viruses, resistance genes from different wild tomato species are introgressed. Recently, the Ty-1 resistance gene was identified, shown to code for an RNA-dependent RNA polymerase and to be allelic with Ty-3. Here we show that upon TYLCV challenging of resistant lines carrying Ty-1 or Ty-3, low virus titers were detected concomitant with the production of relatively high levels of siRNAs whereas, in contrast, susceptible tomato Moneymaker (MM) revealed higher virus titers but lower amounts of siRNAs. Comparative analysis of the spatial genomic siRNA distribution showed a consistent and subtle enrichment for siRNAs derived from the V1 and C3 genes in Ty-1 and Ty-3. In plants containing Ty-2 resistance the virus was hardly detectable, but the siRNA profile resembled the one observed in TYLCV-challenged susceptible tomato (MM). Furthermore, a relative hypermethylation of the TYLCV V1 promoter region was observed in genomic DNA collected from Ty-1 compared with that from (MM). The resistance conferred by Ty-1 was also effective against the bipartite tomato severe rugose begomovirus, where a similar genome hypermethylation of the V1 promoter region was discerned. However, a mixed infection of TYLCV with cucumber mosaic virus compromised the resistance. The results indicate that Ty-1 confers resistance to geminiviruses by increasing cytosine methylation of viral genomes, suggestive of enhanced transcriptional gene silencing. The mechanism of resistance and its durability toward geminiviruses under natural field conditions is discussed.
The evolution of the placenta drives a shift in sexual selection in livebearing fish
Pollux, B.J.A. ; Meredith, R.W. ; Springer, M.S. ; Garland, T. ; Reznick, D.N. - \ 2014
Nature 513 (2014). - ISSN 0028-0836 - p. 233 - 236.
parent-offspring conflict - molecular phylogenetic-relationships - mosquitofish gambusia-holbrooki - reproductive mode - size dimorphism - poeciliidae cyprinodontiformes - maximum-likelihood - viviparity - hypothesis - gene
The evolution of the placenta from a non-placental ancestor causes a shift of maternal investment from pre- to post-fertilization, creating a venue for parent–offspring conflicts during pregnancy1, 2, 3, 4. Theory predicts that the rise of these conflicts should drive a shift from a reliance on pre-copulatory female mate choice to polyandry in conjunction with post-zygotic mechanisms of sexual selection2. This hypothesis has not yet been empirically tested. Here we apply comparative methods to test a key prediction of this hypothesis, which is that the evolution of placentation is associated with reduced pre-copulatory female mate choice. We exploit a unique quality of the livebearing fish family Poeciliidae: placentas have repeatedly evolved or been lost, creating diversity among closely related lineages in the presence or absence of placentation5, 6. We show that post-zygotic maternal provisioning by means of a placenta is associated with the absence of bright coloration, courtship behaviour and exaggerated ornamental display traits in males. Furthermore, we found that males of placental species have smaller bodies and longer genitalia, which facilitate sneak or coercive mating and, hence, circumvents female choice. Moreover, we demonstrate that post-zygotic maternal provisioning correlates with superfetation, a female reproductive adaptation that may result in polyandry through the formation of temporally overlapping, mixed-paternity litters. Our results suggest that the emergence of prenatal conflict during the evolution of the placenta correlates with a suite of phenotypic and behavioural male traits that is associated with a reduced reliance on pre-copulatory female mate choice.
Distribution of anticoagulant rodenticide resistance in Rattus norvegicus in the Netherlands according to Vkorc1 mutations
Meerburg, B.G. ; Gent-Pelzer, M.P.E. van; Schoelitsz, B. ; Lee, T.A.J. van der - \ 2014
Pest Management Science 70 (2014)11. - ISSN 1526-498X - p. 1761 - 1766.
norway rats - tyrosine139cysteine focus - germany - westphalia - chlorophacinone - warfarin - berk. - tests - gene
BACKGROUND Rodenticide resistance to anticoagulants in Rattus norvegicus will lead to increased difficulties in combating these pest animals. Here, the authors present the results of a survey in the Netherlands where tissue samples and droppings were tested using a newly developed TaqMan PCR test for genotypic variation at codon 139 in the Vkorc1 gene associated with anticoagulant rodenticide resistance. Test results are linked to results of a questionnaire that was conducted among pest controllers. RESULTS Genetic mutations at codon 139 of the Vkorc1 gene in R. norvegicus can be encountered in many parts of the Netherlands. In 34/61 rat tails, a genotype was found that is linked to anticoagulant rodenticide resistance (56%). In droppings, 42/169 samples (25%) showed a resistance-mediating genotype. In addition, indications of a clear genetic substructure in the Netherlands were found. In some regions, only resistance-mediating genotypes were found, corroborating results from the questionnaire in which pest controllers indicated they suspected resistance to anticoagulant rodenticides. CONCLUSION This is the first study to demonstrate the presence of multiple genetic mutations at codon 139 of the Vkorc1 gene in R. norvegicus in the Netherlands. As rodenticides should keep their efficacy because they are a last resort in rodent management, more studies are urgently needed that link specific genetic mutations to the efficacy of active substances. © 2014 Society of Chemical Industry
Safety assessment of plant varieties using transcriptomics profiling and a one-class classifier
Dijk, J.P. van; Mello, C.S. de; Voorthuijzen, M.M. ; Hutten, R.C.B. ; Maisonnave Arisi, A.C. ; Jansen, J.J. ; Buydens, L.M.C. ; Voet, H. van der; Kok, E.J. - \ 2014
Regulatory Toxicology and Pharmacology 70 (2014)1. - ISSN 0273-2300 - p. 297 - 303.
potato-tubers - microarray data - risk-assessment - food - identification - components - gene
An important part of the current hazard identification of novel plant varieties is comparative targeted analysis of the novel and reference varieties. Comparative analysis will become much more informative with unbiased analytical approaches, e.g. omics profiling. Data analysis estimating the similarity of new varieties to a reference baseline class of known safe varieties would subsequently greatly facilitate hazard identification. Further biological and eventually toxicological analysis would then only be necessary for varieties that fall outside this reference class. For this purpose, a one-class classifier tool was explored to assess and classify transcriptome profiles of potato (Solanum tuberosum) varieties in a model study. Profiles of six different varieties, two locations of growth, two year of harvest and including biological and technical replication were used to build the model. Two scenarios were applied representing evaluation of a ’different’ variety and a ‘similar’ variety. Within the model higher class distances resulted for the ‘different’ test set compared with the ‘similar’ test set. The present study may contribute to a more global hazard identification of novel plant varieties
Whole-genome sequencing of 234 bulls facilitates mapping of monogenic and complex traits in cattle
Daetwyler, H.D. ; Capitan, A. ; Pausch, H. ; Stothard, P. ; Binsbergen, R. van; Brondum, R.F. ; Liao, X. ; Djari, A. ; Rodriguez, S.C. ; Grohs, C. ; Esquerré, D. ; Bouchez, O. ; Rossignol, M.N. ; Klopp, C. ; Rocha, D. ; Fritz, S. ; Eggen, A. ; Bowman, P.J. ; Coote, D. ; Chamberlain, A.J. ; Anderson, C.L. ; Tassel, C.P. ; Hulsegge, B. ; Goddard, M.E. ; Guldbrandsten, B. ; Lund, M.S. ; Veerkamp, R.F. ; Boichard, D.A. ; Fries, R. ; Hayes, B.J. - \ 2014
Nature Genetics 46 (2014). - ISSN 1061-4036 - p. 858 - 865.
boophilus-microplus resistance - mitotic chromosomes - genotype imputation - holstein calves - dairy-cattle - milk-yield - bos-taurus - condensin - mutations - gene
The 1000 bull genomes project supports the goal of accelerating the rates of genetic gain in domestic cattle while at the same time considering animal health and welfare by providing the annotated sequence variants and genotypes of key ancestor bulls. In the first phase of the 1000 bull genomes project, we sequenced the whole genomes of 234 cattle to an average of 8.3-fold coverage. This sequencing includes data for 129 individuals from the global Holstein-Friesian population, 43 individuals from the Fleckvieh breed and 15 individuals from the Jersey breed. We identified a total of 28.3 million variants, with an average of 1.44 heterozygous sites per kilobase for each individual. We demonstrate the use of this database in identifying a recessive mutation underlying embryonic death and a dominant mutation underlying lethal chrondrodysplasia. We also performed genome-wide association studies for milk production and curly coat, using imputed sequence variants, and identified variants associated with these traits in cattle.
Abscisic acid (ABA) sensitivity regulates desiccation tolerance in germinated Arabidopsis seeds
Maia de Oliveira, J. ; Dekkers, S.J.W. ; Dolle, M. ; Ligterink, W. ; Hilhorst, H.W.M. - \ 2014
New Phytologist 203 (2014)1. - ISSN 0028-646X - p. 81 - 93.
bzip transcription factors - medicago-truncatula seeds - 2c protein phosphatases - signal-transduction - drought tolerance - thaliana - stress - gene - maturation - expression
During germination, orthodox seeds lose their desiccation tolerance (DT) and become sensitive to extreme drying. Yet, DT can be rescued, in a well-defined developmental window, by the application of a mild osmotic stress before dehydration. A role for abscisic acid (ABA) has been implicated in this stress response and in DT re-establishment. However, the path from the sensing of an osmotic cue and its signaling to DT re-establishment is still largely unknown. Analyses of DT, ABA sensitivity, ABA content and gene expression were performed in desiccation- sensitive (DS) and desiccation-tolerant Arabidopsis thaliana seeds. Furthermore, loss and re-establishment of DT in germinated Arabidopsis seeds was studied in ABA-deficient and ABA-insensitive mutants. We demonstrate that the developmental window in which DT can be re-established correlates strongly with the window in which ABA sensitivity is still present. Using ABA biosynthesis and signaling mutants, we show that this hormone plays a key role in DT re-establishment. Surprisingly, re-establishment of DT depends on the modulation of ABA sensitivity rather than enhanced ABA content. In addition, the evaluation of several ABA-insensitive mutants, which can still produce normal desiccation-tolerant seeds, but are impaired in the re-establishment of DT, shows that the acquisition of DT during seed development is genetically different from its re-establishment during germination.
Thermoneutrality results in prominent diet-induced body weight differences in C57BL/6J mice, not paralleled by diet-induced metabolic differences
Hoevenaars, F.P.M. ; Bekkenkamp-Grovenstein, M. ; Janssen, R.J.R.J. ; Heil, S.G. ; Bunschoten, A. ; Hoek-van den Hil, E.F. ; Snaas-Alders, S.H. ; Teerds, K.J. ; Schothorst, E.M. van; Keijer, J. - \ 2014
Molecular Nutrition & Food Research 58 (2014)4. - ISSN 1613-4125 - p. 799 - 807.
adipose-tissue - mitochondrial-function - obesity - fat - thermogenesis - models - health - gene - induction - disease
Scope Mice are usually housed at 20–24°C. At thermoneutrality (28°C) larger diet-induced differences in obesity are seen. We tested whether this leads to large differences in metabolic health parameters. Methods and results We performed a 14-wk dietary intervention in C57BL/6J mice at 28°C and assessed adiposity and metabolic health parameters for a semipurified low fat (10 energy%) diet and a moderate high fat (30 energy%) diet. A large and significant diet-induced differential increase in body weight, adipose tissue mass, adipocyte size, serum leptin level, and, to some extent, cholesterol level was observed. No adipose tissue inflammation was seen. No differential effect of the diets on serum glucose, free fatty acids, triacylglycerides, insulin, adiponectin, resistin, PAI-1, MMP-9, sVCAM-1, sICAM-1, sE-selectin, IL-6, ApoE, fibrinogen levels, or HOMA index was observed. Also in muscle no differential effect on mitochondrial density, mitochondrial respiratory control ratio, or mRNA expression of metabolic genes was found. Finally, in liver no differential effect on weight, triacylglycerides level, aconitase/citrate synthase activity ratio was seen. Conclusion Low fat diet and moderate high fat diet induce prominent body weight differences at thermoneutrality, which is not paralleled by metabolic differences. Our data rather suggest that thermoneutrality alters metabolic homeostasis.
Rearing history affects behaviour and performance of two virulent Nasonovia ribisnigri populations on two lettuce cultivars
Broeke, C.J.M. ten; Dicke, M. ; Loon, J.J.A. van - \ 2014
Entomologia Experimentalis et Applicata 151 (2014)2. - ISSN 0013-8703 - p. 97 - 105.
nematode meloidogyne-incognita - myzus-persicae hemiptera - host-plant resistance - feeding-behavior - tissue localization - aphid resistance - clones - gene - homoptera - biotypes
Many aphid species have become virulent to host-plant resistance, which limits the sustainability of insect resistance breeding. However, when this adaptation to resistant plants is associated with fitness costs for the aphids, virulence can be lost in the absence of resistant plants. For two populations of the lettuce aphid, Nasonovia ribisnigri (Mosely) (Hemiptera: Aphididae), we evaluated whether virulence to Nr-gene-based resistance was lost on a susceptible lettuce, Lactuca sativa L. (Asteraceae), and assessed possible costs of virulence. The feeding behaviour and performance of these aphids, reared and tested on susceptible and resistant lettuce, were investigated. The rearing plant affected feeding behaviour and performance of the aphids. Temporary reduction and long-term loss of virulence were found. The total duration of phloem intake was shorter after being reared on susceptible lettuce and tested on resistant lettuce. In addition, one population had a lower survival on resistant lettuce after being reared on susceptible lettuce. There were also indications of fitness costs of the virulence in both populations.
Mature osteoblasts dedifferentiate in response to traumatic bone injury in the zebrafish fin and skull
Geurtzen, K. ; Knopf, F. ; Wehner, D. ; Huitema, L.F.A. ; Schulte-Merker, S. ; Weidinger, G. - \ 2014
Development 141 (2014). - ISSN 0950-1991 - p. 2225 - 2234.
fibroblast-growth-factor - fracture repair - adult zebrafish - lining cells - regeneration - expression - defects - fate - gene - rats
Zebrafish have an unlimited capacity to regenerate bone after fin amputation. In this process, mature osteoblasts dedifferentiate to osteogenic precursor cells and thus represent an important source of newly forming bone. By contrast, differentiated osteoblasts do not appear to contribute to repair of bone injuries in mammals; rather, osteoblasts form anew from mesenchymal stem cells. This raises the question whether osteoblast dedifferentiation is specific to appendage regeneration, a special feature of the lepidotrichia bone of the fish fin, or a process found more generally in fish bone. Here, we show that dedifferentiation of mature osteoblasts is not restricted to fin regeneration after amputation, but also occurs during repair of zebrafish fin fractures and skull injuries. In both models, mature osteoblasts surrounding the injury downregulate the expression of differentiation markers, upregulate markers of the pre-osteoblast state and become proliferative. Making use of photoconvertible Kaede protein as well as Cre-driven genetic fate mapping, we show that osteoblasts migrate to the site of injury to replace damaged tissue. Our findings suggest a fundamental role for osteoblast dedifferentiation in reparative bone formation in fish and indicate that adult fish osteoblasts display elevated cellular plasticity compared with mammalian bone-forming cells.
Exploring causal networks of bovine milk fatty acids in a multivariate mixed model context
Bouwman, A.C. ; Valente, B.D. ; Janss, L.L.G. ; Bovenhuis, H. ; Rosa, G.J. - \ 2014
Genetics, Selection, Evolution 46 (2014). - ISSN 0999-193X
somatic-cell score - yield - traits - cattle - gene - associations - parameters - cows
BACKGROUND: Knowledge regarding causal relationships among traits is important to understand complex biological systems. Structural equation models (SEM) can be used to quantify the causal relations between traits, which allow prediction of outcomes to interventions applied to such a network. Such models are fitted conditionally on a causal structure among traits, represented by a directed acyclic graph and an Inductive Causation (IC) algorithm can be used to search for causal structures. The aim of this study was to explore the space of causal structures involving bovine milk fatty acids and to select a network supported by data as the structure of a SEM. RESULTS: The IC algorithm adapted to mixed models settings was applied to study 14 correlated bovine milk fatty acids, resulting in an undirected network. The undirected pathway from C4:0 to C12:0 resembled the de novo synthesis pathway of short and medium chain saturated fatty acids. By using prior knowledge, directions were assigned to that part of the network and the resulting structure was used to fit a SEM that led to structural coefficients ranging from 0.85 to 1.05. The deviance information criterion indicated that the SEM was more plausible than the multi-trait model. CONCLUSIONS: The IC algorithm output pointed towards causal relations between the studied traits. This changed the focus from marginal associations between traits to direct relationships, thus towards relationships that may result in changes when external interventions are applied. The causal structure can give more insight into underlying mechanisms and the SEM can predict conditional changes due to such interventions.
Pectic arabinan side chains are essential for pollen cell wall integrity during pollen development
Cankar, K. ; Kortstee, A.J. ; Toonen, M.A.J. ; Wolters-Arts, M. ; Houbein, R. ; Mariani, C. ; Ulvskov, P. ; Jorgensen, B. ; Schols, H.A. ; Visser, R.G.F. ; Trindade, L.M. - \ 2014
Plant Biotechnology Journal 12 (2014)4. - ISSN 1467-7644 - p. 492 - 502.
in-vivo expression - mechanical-properties - potato pectin - arabidopsis - gene - galactan - growth - biosynthesis - mutants - tubers
Pectin is a complex polysaccharide and an integral part of the primary plant cell wall and middle lamella, contributing to cell wall mechanical strength and cell adhesion. To understand the structure–function relationships of pectin in the cell wall, a set of transgenic potato lines with altered pectin composition was analysed. The expression of genes encoding enzymes involved in pectin acetylation, degradation of the rhamnogalacturonan backbone and type and length of neutral side chains, arabinan and galactan in particular, has been altered. Upon crossing of different transgenic lines, some transgenes were not transmitted to the next generation when these lines were used as a pollen donor, suggesting male sterility. Viability of mature pollen was severely decreased in potato lines with reduced pectic arabinan, but not in lines with altered galactan side chains. Anthers and pollen of different developmental stages were microscopically examined to study the phenotype in more detail. Scanning electron microscopy of flowers showed collapsed pollen grains in mature anthers and in earlier stages cytoplasmic protrusions at the site of the of kin pore, eventually leading to bursting of the pollen grain and leaking of the cytoplasm. This phenomenon is only observed after the microspores are released and the tapetum starts to degenerate. Timing of the phenotype indicates a role for pectic arabinan side chains during remodelling of the cell wall when the pollen grain is maturing and dehydrating.
On the relationship between an Asian haplotype on chromosome 6 that reduces androstenone levels in boars and the differential expression of SULT2A1 in the testis
Hidalgo, A.M. ; Bastiaansen, J.W.M. ; Harlizius, B. ; Megens, H.J.W.C. ; Madsen, O. ; Crooijmans, R.P.M.A. ; Groenen, M.A.M. - \ 2014
BMC Genetics 15 (2014). - ISSN 1471-2156
genome-wide association - quantitative trait loci - 16-androstene steroids - pig genomes - taint - gene - domestication - reveals - sulfoconjugation - visualization
Background Androstenone is one of the major compounds responsible for boar taint, a pronounced urine-like odor produced when cooking boar meat. Several studies have identified quantitative trait loci (QTL) for androstenone level on Sus scrofa chromosome (SSC) 6. For one of the candidate genes in the region SULT2A1, a difference in expression levels in the testis has been shown at the protein and RNA level. Results Haplotypes were predicted for the QTL region and their effects were estimated showing that haplotype 1 was consistently related with a lower level, and haplotype 2 with a higher level of androstenone. A recombinant haplotype allowed us to narrow down the QTL region from 3.75 Mbp to 1.94 Mbp. An RNA-seq analysis of the liver and testis revealed six genes that were differentially expressed between homozygotes of haplotypes 1 and 2. Genomic sequences of these differentially expressed genes were checked for variations within potential regulatory regions. We identified one variant located within a CpG island that could affect expression of SULT2A1 gene. An allele-specific expression analysis in the testis did not show differential expression between the alleles of SULT2A1 located on the different haplotypes in heterozygous animals. However a synonymous mutation C166T (SSC6: 49,117,861 bp in Sscrofa 10.2; C/T) was identified within the exon 2 of SULT2A1 for which the haplotype 2 only had the C allele which was higher expressed than the T allele, indicating haplotype-independent allelic-imbalanced expression between the two alleles. A phylogenetic analysis for the 1.94 Mbp region revealed that haplotype 1, associated with low androstenone level, originated from Asia. Conclusions Differential expression could be observed for six genes by RNA-seq analysis. No difference in the ratio of C:T expression of SULT2A1 for the haplotypes was found by the allele-specific expression analysis, however, a difference in expression between the C over T allele was found for a variation within SULT2A1, showing that the difference in androstenone levels between the haplotypes is not caused by the SNP in exon 2.
Hyperactivity and tree-top disease induced by the baculovirus AcMNPV in Spodoptera exigua larvae are governed by independent mechanisms
Houte, S. van; Ros, V.I.D. ; Oers, M.M. van - \ 2014
Naturwissenschaften 101 (2014)4. - ISSN 0028-1042 - p. 347 - 350.
behavior - host - gene
Although many parasites are known to manipulate the behavior of their hosts, the mechanisms underlying such manipulations are largely unknown. Baculoviruses manipulate the behavior of caterpillar hosts by inducing hyperactivity and by inducing climbing behavior leading to death at elevated positions (tree-top disease or Wipfelkrankheit). Whether hyperactivity and tree-top disease are independent manipulative strategies of the virus is unclear. Recently, we demonstrated the involvement of the protein tyrosine phosphatase (ptp) gene of the baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV) in the induction of hyperactivity in Spodoptera exigua larvae. Here we show that AcMNPV ptp is not required for tree-top disease, indicating that in S. exigua baculovirus-induced hyperactivity and tree-top disease are independently induced behaviors that are governed by distinct mechanisms.
Single nucleus genome sequencing reveals high similarity among nuclei of an endomycorrhizal fungus
Lin, K. ; Limpens, E.H.M. ; Zhang, Z. ; Ivanov, S. ; Saunders, D.G.O. ; Mu, D. ; Pang, E. ; Cao, H. ; Cha, H. ; Lin, T. ; Zhou, Q. ; Shang, Y. ; Li, Y. ; Sharma, T.C. ; Velzen, R. van; Ruijter, N.C.A. de; Aanen, D.K. ; Win, J. ; Kamoun, S. ; Bisseling, T. ; Geurts, R. ; Huang, S.W. - \ 2014
Plos Genetics 10 (2014)1. - ISSN 1553-7404 - 13 p.
arbuscular mycorrhizal fungi - pathogen phytophthora-infestans - glomus-intraradices - sexual reproduction - protein families - cdna sequences - kingdom fungi - gene - identification - efficient
Nuclei of arbuscular endomycorrhizal fungi have been described as highly diverse due to their asexual nature and absence of a single cell stage with only one nucleus. This has raised fundamental questions concerning speciation, selection and transmission of the genetic make-up to next generations. Although this concept has become textbook knowledge, it is only based on studying a few loci, including 45S rDNA. To provide a more comprehensive insight into the genetic makeup of arbuscular endomycorrhizal fungi, we applied de novo genome sequencing of individual nuclei of Rhizophagus irregularis. This revealed a surprisingly low level of polymorphism between nuclei. In contrast, within a nucleus, the 45S rDNA repeat unit turned out to be highly diverged. This finding demystifies a long-lasting hypothesis on the complex genetic makeup of arbuscular endomycorrhizal fungi. Subsequent genome assembly resulted in the first draft reference genome sequence of an arbuscular endomycorrhizal fungus. Its length is 141 Mbps, representing over 27,000 protein-coding gene models. We used the genomic sequence to reinvestigate the phylogenetic relationships of Rhizophagus irregularis with other fungal phyla. This unambiguously demonstrated that Glomeromycota are more closely related to Mucoromycotina than to its postulated sister Dikarya.
Diversity of Global Rice Markets and the Science Required for Consumer-Targeted Rice Breeding
Calingacion, M.N. ; Laborte, A.G. ; Nelson, A. ; Resurreccion, A. ; Chrystal Concepcion, J. ; Dara Daygon, V. ; Mumm, R. ; Reinke, R. ; Dipti, S. ; Zaczuk Bassinello, P. ; Manful, J. ; Sophany, S. ; Cordero Lara, K. ; Bao, J. ; Xie, L. ; Loaiza, K. ; El-hissewy, A. ; Gayin, J. ; Sharma, N. ; Rajeswari, S. ; Manonmani, S. ; Shobha Rani, N. ; Kota, S. ; Dewi Indrasari, S. ; Habibi, F. ; Hosseini, M. ; Tavasoli, F. ; Suzuki, K. ; Umemoto, T. ; Boualaphanh, C. ; Hong Lee, H. ; Pang Hung, Y. ; Ramli, A. ; Pa Aung, P. ; Ahmad, R. ; Iqbal Wattoo, J. ; Bandonill, E. ; Romero, M. ; Moita Brites, C. ; Hafeel, R. ; Sheng Lur, H. ; Cheaupun, K. ; Jongdee, S. ; Blanco, P. ; Bryant, R. ; Thi Lang, N. ; Hall, R.D. - \ 2014
PLoS ONE 9 (2014)1. - ISSN 1932-6203 - 12 p.
single-nucleotide polymorphisms - oryza-sativa l. - starch-synthase-iia - grain length - gelatinization temperature - gel consistency - eating quality - gene - gs3 - association
With the ever-increasing global demand for high quality rice in both local production regions and with Western consumers, we have a strong desire to understand better the importance of the different traits that make up the quality of the rice grain and obtain a full picture of rice quality demographics. Rice is by no means a ‘one size fits all’ crop. Regional preferences are not only striking, they drive the market and hence are of major economic importance in any rice breeding / improvement strategy. In this analysis, we have engaged local experts across the world to perform a full assessment of all the major rice quality trait characteristics and importantly, to determine how these are combined in the most preferred varieties for each of their regions. Physical as well as biochemical characteristics have been monitored and this has resulted in the identification of no less than 18 quality trait combinations. This complexity immediately reveals the extent of the specificity of consumer preference. Nevertheless, further assessment of these combinations at the variety level reveals that several groups still comprise varieties which consumers can readily identify as being different. This emphasises the shortcomings in the current tools we have available to assess rice quality and raises the issue of how we might correct for this in the future. Only with additional tools and research will we be able to define directed strategies for rice breeding which are able to combine important agronomic features with the demands of local consumers for specific quality attributes and hence, design new, improved crop varieties which will be awarded success in the global market.
Expression of the Aspergillus terreus itaconic acid biosynthesis cluster in Aspergillus niger
Straat, L. van der; Vernooij, M. ; Lammers, M. ; Berg, W.A.M. van den; Schonewille, T. ; Cordewener, J. ; Meer, I. van der; Koops, A.J. ; Graaff, L.H. de - \ 2014
Microbial Cell Factories 13 (2014). - ISSN 1475-2859 - 9 p.
BACKGROUND: Aspergillus terreus is a natural producer of itaconic acid and is currently used to produce itaconic acid on an industrial scale. The metabolic process for itaconic acid biosynthesis is very similar to the production of citric acid in Aspergillus niger. However, a key enzyme in A. niger, cis-aconitate decarboxylase, is missing. The introduction of the A. terreus cadA gene in A. niger exploits the high level of citric acid production (over 200 g per liter) and theoretically can lead to production levels of over 135 g per liter of itaconic acid in A. niger. Given the potential for higher production levels in A. niger, production of itaconic acid in this host was investigated. RESULTS: Expression of Aspergillus terreus cis-aconitate decarboxylase in Aspergillus niger resulted in the production of a low concentration (0.05 g/L) of itaconic acid. Overexpression of codon-optimized genes for cis-aconitate decarboxylase, a mitochondrial transporter and a plasma membrane transporter in an oxaloacetate hydrolase and glucose oxidase deficient A. niger strain led to highly increased yields and itaconic acid production titers. At these higher production titers, the effect of the mitochondrial and plasma membrane transporters was much more pronounced, with levels being 5-8 times higher than previously described. CONCLUSIONS: Itaconic acid can be produced in A. niger by the introduction of the A. terreus cis-aconitate decarboxylase encoding cadA gene. This results in a low itaconic acid production level, which can be increased by codon-optimization of the cadA gene for A. niger. A second crucial requirement for efficient production of itaconic acid is the expression of the A. terreus mttA gene, encoding a putative mitochondrial transporter. Expression of this transporter results in a twenty-fold increase in the secretion of itaconic acid. Expression of the A. terreus itaconic acid cluster consisting of the cadA gene, the mttA gene and the mfsA gene results in A. niger strains that produce over twenty five-fold higher levels of itaconic acid and show a twenty-fold increase in yield compared to a strain expressing only CadA
Bioactivity screening and mass spectrometric confirmation for the detection of PPAR-delta agonists that increase type 1 muscle fibres
Bovee, T.F.H. ; Blokland, M.H. ; Kersten, A.H. ; Hamers, A.R.M. ; Heskamp, H.H. ; Essers, M.L. ; Nielen, M.W.F. ; Ginkel, L.A. van - \ 2014
Analytical and Bioanalytical Chemistry 406 (2014). - ISSN 1618-2642 - p. 705 - 713.
human skeletal-muscle - gamma - macrophages - expression - receptors - cells - acid - gene - fat
Sensitive and robust bioassays able to detect nuclear receptor activation are very useful for veterinary and doping control, pharmaceutical industry and environmental scientists. Here, we used bioassays based on human leukemic monocyte lymphoma U937 and human liver hepatocellular carcinoma HepG2 cell lines to detect the ligand-induced activation of the peroxisome proliferator-activated receptor delta (PPARd). Exposure of U937 cells to the PPARd agonist GW501516 resulted in a marked increase in mRNA expression of the PPARd target gene Angptl4 which was quantified by qRTPCR analysis. Exposure ofHepG2 cells transiently transfected with a PPARd expression plasmid and a PPAR-response element-driven luciferase reporter plasmid to PPARd agonists GW501516, GW610742 and L-165041 resulted in clear dose–response curves. Although the qRT-PCR resulted in higher fold inductions, the luciferase assay with transfected HepG2 cells is cheaper and quicker and about ten times more sensitive to GW501516 compared to analysis of Angptl4 mRNA expression in U937 cells by qRT-PCR. The HepG2- based luciferase assay was therefore used to screen GW501516-spiked supplements and feed and water samples. After liquid extraction and clean-up by solid phase extraction using a weak anion exchange column, extracts were screened in the HepG2 bioassay followed by confirmation with a newly developed UPLC-MS/MS method, using two transitions for each compound, i.e., for GW501516, 454.07>188.15 (collision energy (CE) 46 V) and 454.07>257.08 (CE 30 V); for GW610742, 472.07>206.2 (CE 48 V) and 472.07>275.08 (CE 30 V); and for L-165041, 401.2>193.15 (CE 26 V) and 401.2>343.2 (CE 20 V).
Anaerostipes rhamnosivorans sp. nov., a human intestinal butyrate forming bacterium
Bui, T.P.N. ; Vos, W.M. de; Plugge, C.M. - \ 2014
International Journal of Systematic and Evolutionary Microbiology 64 (2014)3. - ISSN 1466-5026 - p. 787 - 793.
renaturation rates - dna hybridization - human colon - human feces - microbiota - gene - gut
A novel butyrate producing bacterium, strain 1y-2T, was isolated from a stool sample of a 1-year old, healthy Dutch infant. The isolate was obtained by using lactate and acetate as sources of carbon and energy. The strain was Gram-variable, strictly anaerobic, spore-forming and formed curly rod-shaped cells that fermented glucose into butyrate, lactate, formate and acetate as main products. The G + C composition of the strain was 44.5 % and its major cellular fatty acids were C12:00, C19:1 iso I and C16:00. Strain 1y-2T was related to Anaerostipes caccae DSM14662T based on 16S rRNA analysis with 3 % divergence but hybridization studies of their genomic DNA revealed only 33 % similarity. Moreover, strain 1y-2T showed marked physiological and biochemical differences with known Anaerostipes species. Based on the phylogenetic, chemotypic and phenotypic criteria, we propose that strain 1y-2T should be classified in the genus Anaerostipes as a novel species, Anaerostipes rhamnosivorans sp. nov. The type strain is 1y-2T (= DSM 26241T = KCTC 15316T)
Molecular characterization and functional analyses of ZtWor1, a transcriptional regulator of the fungal wheat pathogen Zymoseptoria tritici
Mirzadi Gohari, A. ; Mehrabi, R. ; Robert, O. ; Ince, I.A. ; Boeren, J.A. ; Schuster, M. ; Steinberg, G. ; Wit, P.J.G.M. de; Kema, G.H.J. - \ 2014
Molecular Plant Pathology 15 (2014)4. - ISSN 1464-6722 - p. 394 - 405.
phytopathogen mycosphaerella-graminicola - candida-albicans - sample preparation - septoria-tritici - azole fungicides - master regulator - gene - protein - resistance - cultivars
Zymoseptoria tritici causes the major fungal wheat disease septoria tritici blotch, and is increasingly being used as a model for transmission and population genetics, as well as host–pathogen interactions. Here, we study the biological function of ZtWor1, the orthologue of Wor1 in the fungal human pathogen Candida albicans, as a representative of a superfamily of regulatory proteins involved in dimorphic switching. In Z.¿tritici, this gene is pivotal for pathogenesis, as ZtWor1 mutants were nonpathogenic and complementation restored the wild-type phenotypes. In¿planta expression analyses showed that ZtWor1 is up-regulated during the initiation of colonization and fructification, and regulates candidate effector genes, including one that was discovered after comparative proteome analysis of the Z.¿tritici wild-type strain and the ZtWor1 mutant, which was particularly expressed in¿planta. Cell fusion and anastomosis occur frequently in ZtWor1 mutants, reminiscent of mutants of MgGpb1, the ß-subunit of the heterotrimeric G protein. Comparative expression of ZtWor1 in knock-out strains of MgGpb1 and MgTpk2, the catalytic subunit of protein kinase A, suggests that ZtWor1 is downstream of the cyclic adenosine monophosphate (cAMP) pathway that is crucial for pathogenesis in many fungal plant pathogens
Pseudogenization in pathogenic fungi with different host plants and lifestyles might reflect their evolutionary past
Burgt, I.A. van der; Karimi, M. ; Bahkali, A.H. ; Wit, P.J.G.M. de - \ 2014
Molecular Plant Pathology 15 (2014)2. - ISSN 1464-6722 - p. 133 - 144.
cladosporium-fulvum - gene - resistance - prediction - software - proteins - genomics - update - locus
Pseudogenes are genes with significant homology to functional genes but contain disruptive mutations (DMs) leading to production of non- or partially functional proteins. Little is known about pseudogenization in pathogenic fungi with different lifestyles. Here we report on identification of DMs causing pseudogenes in the genomes of the fungal plant pathogens Botrytis cinerea, Cladosporium fulvum, Dothistroma septosporum, Mycosphaerella fijiensis, Verticillium dahliae and Zymoseptoria tritici. In these fungi we have identified 1740 gene models containing 2795 DMs obtained by an alignment-based gene prediction method. The contribution of sequencing errors to DMs was minimized by analyses of resequenced genomes to obtain a refined data set of 924 gene models containing 1666 true DMs. The frequency of pseudogenes varied from 1 to 5% in the gene catalogues of these fungi, being the highest in the asexually reproducing fungi C. fulvum (4.9%), followed by D. septosporum (2.4%) and V. dahliae (2.1%). The majority of pseudogenes does not represent recent gene duplications, but members of multi-gene families and unitary genes. In general there was no bias for pseudogenization of specific genes in the six fungi. Single exceptions are those encoding secreted proteins including proteases which appeared more frequently pseudogenized in C. fulvum than in D. septosporum. Most pseudogenes present in these two phylogenically closely related fungi are not shared suggesting that they are related to adaptation to a different host (tomato versus pine) and lifestyle (biotroph versus hemi-biotroph)
Chaperones of the endoplasmic reticulum are required for Ve1-mediated resistance to Verticillium
Liebrand, T.W.H. ; Kombrink, A. ; Zhang, Z. ; Sklenar, J. ; Jones, A.M.E. ; Robatzek, S. ; Thomma, B.P.H.J. ; Joosten, M.H.A.J. - \ 2014
Molecular Plant Pathology 15 (2014)1. - ISSN 1464-6722 - p. 109 - 117.
receptor-like proteins - defective brassinosteroid receptor - pattern-recognition receptors - er quality-control - plant innate immunity - cell-surface - tomato ve1 - arabidopsis - gene - perception
The tomato receptor-like protein (RLP) Ve1 mediates resistance to the vascular fungal pathogen Verticillium dahliae. To identify the proteins required for Ve1 function, we transiently expressed and immunopurified functional Ve1-enhanced green fluorescent protein (eGFP) from Nicotiana benthamiana leaves, followed by mass spectrometry. This resulted in the identification of peptides originating from the endoplasmic reticulum (ER)-resident chaperones HSP70 binding proteins (BiPs) and a lectin-type calreticulin (CRT). Knock-down of the different BiPs and CRTs in tomato resulted in compromised Ve1-mediated resistance to V.dahliae in most cases, showing that these chaperones play an important role in Ve1 functionality. Recently, it has been shown that one particular CRT is required for the biogenesis of the RLP-type Cladosporium fulvum resistance protein Cf-4 of tomato, as silencing of CRT3a resulted in a reduced pool of complex glycosylated Cf-4 protein. In contrast, knock-down of the various CRTs in N.benthamiana or N.tabacum did not result in reduced accumulation of mature complex glycosylated Ve1 protein. Together, this study shows that the BiP and CRT ER chaperones differentially contribute to Cf-4- and Ve1-mediated immunity.