Characterization of polarity development through 2- and 3-D imaging during the initial phase of microspore embryogenesis in Brassica napus L.
Dubas, E. ; Custers, J. ; Kieft, H. ; Wedzony, M. ; Lammeren, A.A.M. van - \ 2014
Protoplasma 251 (2014)1. - ISSN 0033-183X - p. 103 - 113.
nuclear-dna synthesis - cv topas - extracellular-matrix - sporophytic development - cultured microspores - actin-filament - heat-shock - in-vitro - induction - pollen
Isolated microspores of B. napus in culture change their developmental pathway from gametophytic to sporophytic and form embryo-like structures (ELS) upon prolonged heat shock treatment (5 days at 32 °C). ELS express polarity during the initial days of endosporic development. In this study, we focussed on the analysis of polarity development of ELS without suspensor. Fluorescence microscopy and 3-D confocal laser scanning microscopy (CLSM) without tissue interfering enabled us to get a good insight in the distribution of nuclei, mitochondria and endoplasmic reticulum(ER), the architecture of microtubular (MT) cytoskeleton and the places of 5-bromo- 2'-deoxy-uridine (BrdU) incorporation in successive stages of microspore embryogenesis. Scanning electron microscopy (SEM) analysis revealed, for the first time, the appearance of a fibrillar extracellular matrix-like structure (ECM-like structure) in androgenic embryos without suspensor. Two types of endosporic development were distinguished based upon the initial location of the microspore nucleus. The polarity of dividing and growing cells was recognized by the differential distributions of organelles, by the organization of the MT cytoskeleton and by the visualization of DNA synthesis in the cell cycle. The directional location of nuclei, ER, mitochondria and starch grains in relation to theMTs configurations were early polarity indicators. Both exine rupture and ECMlike structure on the outer surfaces of ELS are supposed to stabilize ELS's morphological polarity. As the role of cell polarity during early endosporic microspore embryogenesis in apical–basal cell fate determination remains unclear, microspore culture system provides a powerful in vitro tool for studying the developmental processes that take place during the earliest stages of plant embryogenesis.
Genetic variation for stress-response hormesis in C. elegans lifespan
Rodriguez Sanchez, M. ; Snoek, L.B. ; Riksen, J.A.G. ; Bevers, R.P.J. ; Kammenga, J.E. - \ 2012
Experimental Gerontology 47 (2012)8. - ISSN 0531-5565 - p. 581 - 587.
quantitative trait loci - genotype-environment interactions - nematode caenorhabditis-elegans - long-lived mutant - drosophila-melanogaster - heat-shock - history traits - natural variation - longevity - resistance
Increased lifespan can be associated with greater resistance to many different stressors, most notably thermal stress. Such hormetic effects have also been found in C. elegans where short-term exposure to heat lengthens the lifespan. Genetic investigations have been carried out using mutation perturbations in a single genotype, the wild type Bristol N2. Yet, induced mutations do not yield insight regarding the natural genetic variation of thermal tolerance and lifespan. We investigated the genetic variation of heat-shock recovery, i.e. hormetic effects on lifespan and associated quantitative trait loci (QTL) in C. elegans. Heat-shock resulted in an 18% lifespan increase in wild type CB4856 whereas N2 did not show a lifespan elongation. Using recombinant inbred lines (RILs) derived from a cross between wild types N2 and CB4856 we found natural variation in stress-response hormesis in lifespan. Approx. 28% of the RILs displayed a hormesis effect in lifespan. We did not find any hormesis effects for total offspring. Across the RILs there was no relation between lifespan and offspring. The ability to recover from heat-shock mapped to a significant QTL on chromosome II which overlapped with a QTL for offspring under heat-shock conditions. The QTL was confirmed by introgressing relatively small CB4856 regions into chromosome II of N2. Our observations show that there is natural variation in hormetic effects on C. elegans lifespan for heat-shock and that this variation is genetically determined.
Effect of acid stress on protein expression and phosphorylation in Lactobacillus rhamnosus GG
Koponen, J. ; Laakso, K. ; Koskenniemi, K. ; Kankainen, M. ; Savijoki, K. ; Nyman, T.A. ; Vos, W.M. de; Tynkkynen, S. ; Kalkkinen, N. ; Varmanen, P. - \ 2012
Journal of Proteomics 75 (2012)4. - ISSN 1874-3919 - p. 1357 - 1374.
gram-positive bacteria - low-ph - lactococcus-lactis - proteomic analysis - bacillus-subtilis - heat-shock - ser/thr/tyr phosphoproteome - tolerance response - probiotic bacteria - mass-spectrometry
Acidic environments encountered in food products and during gastrointestinal tract passage affect the survival of bacteria that are marketed as probiotics. In this study, the global proteome responses of the probiotic lactic acid bacterium Lactobacillus rhamnosus GG to two physiologically relevant pH conditions (pH 4.8 and pH 5.8) were studied by 2-D DIGE. The proteomics data were complemented with transcriptome analyses by whole-genome DNA microarrays. The cells were cultured in industrial-type whey medium under strictly defined bioreactor conditions. In total, 2-D DIGE revealed the pH-dependent formation of 92 protein spots. In response to lower pH conditions, the strongest up-regulation of all proteins was detected for a predicted surface antigen, LGG_02016. In addition, the acid pH was found to up-regulate the expression of F(0)F(1)-ATP synthase genes whereas the abundance of proteins participating in nucleotide biosynthesis and protein synthesis was significantly diminished. Moreover, the results suggest that L. rhamnosus GG modulates its pyruvate metabolism depending on the growth pH. Furthermore, a growth pH-dependent protein phosphorylation phenomenon was detected in several L. rhamnosus GG proteins with ProQ Diamond 2-DE gel staining. Proteins participating in central cellular pathways were shown to be phosphorylated, and the phosphorylation of glycolytic enzymes was found to be especially extensive
Microtubule configurations and nuclear DNA synthesis during initiation of suspensor-bearing embryos from Brassica napus cv. Topas microspores
Dubas, E. ; Custers, J.B.M. ; Kieft, H. ; Wedzony, M. ; Lammeren, A.A.M. van - \ 2011
Plant Cell Reports 30 (2011)11. - ISSN 0721-7714 - p. 2105 - 2116.
heat-shock - sporophytic development - cultured microspores - rape embryogenesis - in-vitro - induction - pollen - organization - division - visualization
In the new Brassica napus microspore culture system, wherein embryos with suspensors are formed, ab initio mimics zygotic embryogenesis. The system provides a powerful in vitro tool for studying the diverse developmental processes that take place during early stages of plant embryogenesis. Here, we studied in this new culture system both the temporal and spatial distribution of nuclear DNA synthesis places and the organization of the microtubular (MT) cytoskeleton, which were visualized with a refined whole mount immunolocalization technology and 3D confocal laser scanning microscopy. A 'mild' heat stress induced microspores to elongate, to rearrange their MT cytoskeleton and to re-enter the cell cycle and perform a predictable sequence of divisions. These events led to the formation of a filamentous suspensor-like structure, of which the distal tip cell gave rise to the embryo proper. Cells of the developing pro-embryo characterized endoplasmic (EMTs) and cortical microtubules (CMTs) in various configurations in the successive stages of the cell cycle. However, the most prominent changes in MT configurations and nuclear DNA replication concerned the first sporophytic division occurring within microspores and the apical cell of the pro-embryo. Microspore embryogenesis was preceded by pre-prophase band formation and DNA synthesis. The apical cell of the proembryo exhibited a random organization of CMTs and, in relation to this, isotropic expansion occurred, mimicking the development of the apical cell of the zygotic situation. Moreover, the apical cell entered the S phase shortly before it divided transversally at the stage that the suspensor was 3-8 celled.
Bacterial SOS response: a food safety perspective
Veen, S. van der; Abee, T. - \ 2011
Current Opinion in Biotechnology 22 (2011)2. - ISSN 0958-1669 - p. 136 - 142.
monocytogenes egd-e - escherichia-coli - listeria-monocytogenes - dna-damage - lactococcus-lactis - reca protein - heat-shock - antibiotic ciprofloxacin - staphylococcus-aureus - biofilm communities
The SOS response is a conserved inducible pathway in bacteria that is involved in DNA repair and restart of stalled replication forks. Activation of the SOS response can result in stress resistance and mutagenesis. In food processing facilities and during food preservation, bacteria are exposed to stresses and stimuli that potentially activate the SOS response, resulting in resistant or adapted bacteria. This review places the bacterial SOS response in a food safety perspective by providing an overview of the known triggers of the SOS response mechanism and its impact on the survival of spoilage and pathogenic bacteria.
Global transcriptomics analysis of the Desulfovibrio vulgaris change from syntrophic growth with Methanosarcina barkeri to sulfidogenic metabolism
Plugge, C.M. ; Scholten, J.C.M. ; Culley, D.E. ; Nie, L. ; Brockman, F.J. ; Zhang, W. - \ 2010
Microbiology 156 (2010)9. - ISSN 1350-0872 - p. 2746 - 2756.
sulfate-reducing bacteria - gene-expression analysis - electron-transfer - methane production - heat-shock - oligonucleotide microarrays - vulgatis hildenborough - anaerobic-bacteria - mass-spectrometry - oxidative stress
Desulfovibrio vulgaris is a metabolically flexible micro-organism. It can use sulfate as an electron acceptor to catabolize a variety of substrates, or in the absence of sulfate can utilize organic acids and alcohols by forming a syntrophic association with a hydrogen-scavenging partner to relieve inhibition by hydrogen. These alternative metabolic types increase the chance of survival for D. vulgaris in environments where one of the potential external electron acceptors becomes depleted. In this work, whole-genome D. vulgaris microarrays were used to determine relative transcript levels as D. vulgaris shifted its metabolism from syntrophic in a lactate-oxidizing dual-culture with Methanosarcina barkeri to a sulfidogenic metabolism. Syntrophic dual-cultures were grown in two independent chemostats and perturbation was introduced after six volume changes with the addition of sulfate. The results showed that 132 genes were differentially expressed in D. vulgaris 2 h after addition of sulfate. Functional analyses suggested that genes involved in cell envelope and energy metabolism were the most regulated when comparing syntrophic and sulfidogenic metabolism. Upregulation was observed for genes encoding ATPase and the membrane-integrated energy-conserving hydrogenase (Ech) when cells shifted to a sulfidogenic metabolism. A five-gene cluster encoding several lipoproteins and membrane-bound proteins was downregulated when cells were shifted to a sulfidogenic metabolism. Interestingly, this gene cluster has orthologues found only in another syntrophic bacterium, Syntrophobacter fumaroxidans, and four recently sequenced Desulfovibrio strains. This study also identified several novel c-type cytochrome-encoding genes, which may be involved in the sulfidogenic metabolism
Genome-wide screen for Listeria monocytogenes genes important for growth at high temperatures
Veen, S. van der; Abee, T. ; Vos, W.M. de; Wells-Bennik, M.H.J. - \ 2009
FEMS Microbiology Letters 295 (2009)2. - ISSN 0378-1097 - p. 195 - 203.
staphylococcus-aureus - bacillus-subtilis - stress tolerance - escherichia-coli - cell-division - heat-shock - expression - virulence - identification - mutants
Listeria monocytogenes is a Gram-positive food-borne pathogen that is able to grow over a wide temperature range. Although the class I and class III heat-shock genes are known to play an important role in heat shock, information on genes that are essential for growth at high temperatures is scarce. To determine which genes are important for growth at high temperatures (42.5-43 degrees C), we performed a random insertion screening in L. monocytogenes, rendering 28 temperature-sensitive mutants. These mutants showed insertions in genes that play a role in transcription regulation, cell-wall biosynthesis, cell division, translation, transport, sensing, and specific stress responses like the SOS response and the class III heat-shock response. Some of these mutants showed altered morphological characteristics such as cell elongation, reduced cell length, or sickle shapes. Furthermore, the majority of the mutants showed increased heat inactivation after exposure to 55 degrees C compared with the wild-type strain. The role of the specific genes in relation to growth at high temperatures is discussed
The growth limits of a large number of Listeria monocytogenes strains at combinations of stresses show serotype- and niche-specific traits
Veen, S. van der; Moezelaar, R. ; Abee, T. ; Wells-Bennik, M.H.J. - \ 2008
Journal of Applied Microbiology 105 (2008)5. - ISSN 1364-5072 - p. 1246 - 1258.
osmolyte transporters betl - food-borne pathogen - comparative genomics - dairy-products - water activity - inoculum size - lactic-acid - heat-shock - scott-a - low ph
Aims: The aim of this study was to associate the growth limits of Listeria monocytogenes during exposure to combined stresses with specific serotypes or origins of isolation, and identify potential genetic markers. Methods and Results: The growth of 138 strains was assessed at different temperatures using combinations of low pH, sodium lactate, and high salt concentrations in brain heart infusion broth. None of the strains was able to grow at pH ¿ 4·4, aw ¿ 0·92, or pH ¿ 5·0 combined with aw ¿ 0·94. In addition, none of the strains grew at pH ¿ 5·2 and NaLac ¿ 2%. At 30°C, the serotype 4b strains showed the highest tolerance to low pH and high NaCl concentrations at both pH neutral (pH 7·4) and mild acidic conditions (pH 5·5). At 7°C, the serotype 1/2b strains showed the highest tolerance to high NaCl concentrations at both pH 7·4 and 5·5. Serotype 1/2b meat isolates showed the highest tolerance to low pH in the presence of 2% sodium lactate at 7°C. ORF2110 and gadD1T1 were identified as potential biomarkers for phenotypic differences. Conclusions: Differences in growth limits were identified between specific L. monocytogenes strains and serotypes, which could in some cases be associated with specific genetic markers. Significance and Impact of the Study: Our data confirm the growth limits of L. monocytogenes as set out by the European Union for ready-to-eat foods and provides an additional criterion. The association of L. monocytogenes serotypes with certain stress responses might explain the abundance of certain serotypes in retail foods while others are common in clinical cases.
Functional ecological genomics to demonstrate general and specific responses to abiotic stress
Roelofs, D. ; Aarts, M.G.M. ; Schat, H. ; Straalen, N.M. van - \ 2008
Functional Ecology 22 (2008)1. - ISSN 0269-8463 - p. 8 - 18.
hyperaccumulator thlaspi-caerulescens - springtail orchesella-cincta - messenger-rna expression - length cdna microarray - vulgaris moench garcke - high-salinity stresses - caenorhabditis-elegans - arabidopsis-thaliana - transcription factor - heat-shock
1. Stress is a major component of natural selection in soil ecosystems. The most prominent abiotic stress factors in the field are temperature extremes (heat, cold), dehydration (drought), high salinity and specific toxic compounds such as heavy metals. Organisms are able to deal with these stresses to a certain extent, which determines the limits of their ecological amplitudes. Functional genomic tools are now becoming available to study stress in ecologically relevant soil organisms. 2. Here we give an overview of transcriptomic studies aiming to elucidate how plants and soil invertebrates respond and adapt to a stressful environment. The picture emerging from signalling pathways and transcription factors identified in transcription profiling studies suggests that there is a large overlap of genomic responses to drought, salinity and cold; however, heat and heavy metals trigger different stress response pathways. 3. The heat shock response and the oxidative stress response seem to represent universal components of the environmental stress response (ESR). Furthermore, the commonality across plants and animals seems to be higher in effector genes than in transcriptional regulators. 4. Finally, adaptation to stress factors in soil seems to evolve through enhanced constitutive transcription of otherwise stress responsive genes both in plants and animals
Quantification of the Effects of Salt Stress and Physiological State on Thermotolerance of Bacillus cereus ATCC 10987 and ATCC 14579
Besten, H.M.W. den; Mataragas, M. ; Moezelaar, R. ; Abee, T. ; Zwietering, M.H. - \ 2006
Applied and Environmental Microbiology 72 (2006)9. - ISSN 0099-2240 - p. 5884 - 5894.
shock-induced thermotolerance - listeria-monocytogenes - heat-shock - thermal inactivation - bacterial-spores - survival curves - growth-phase - models - temperature - proteins
The food-borne pathogen Bacillus cereus can acquire enhanced thermal resistance through multiple mechanisms. Two Bacillus cereus strains, ATCC 10987 and ATCC 14579, were used to quantify the effects of salt stress and physiological state on thermotolerance. Cultures were exposed to increasing concentrations of sodium chloride for 30 min, after which their thermotolerance was assessed at 50°C. Linear and nonlinear microbial survival models, which cover a wide range of known inactivation curvatures for vegetative cells, were fitted to the inactivation data and evaluated. Based on statistical indices and model characteristics, biphasic models with a shoulder were selected and used for quantification. Each model parameter reflected a survival characteristic, and both models were flexible, allowing a reduction of parameters when certain phenomena were not present. Both strains showed enhanced thermotolerance after preexposure to (non)lethal salt stress conditions in the exponential phase. The maximum adaptive stress response due to salt preexposure demonstrated for exponential-phase cells was comparable to the effect of physiological state on thermotolerance in both strains. However, the adaptive salt stress response was less pronounced for transition- and stationary-phase cells. The distinct tailing of strain ATCC 10987 was attributed to the presence of a subpopulation of spores. The existence of a stable heat-resistant subpopulation of vegetative cells could not be demonstrated for either of the strains. Quantification of the adaptive stress response might be instrumental in understanding adaptation mechanisms and will allow the food industry to develop more accurate and reliable stress-integrated predictive modeling to optimize minimal processing conditions.
PCIB an antiauxin enhances microspore embryogenisis in microspore culture of Brassica juncea
Agarwal, P.K. ; Agarwal, P. ; Custers, J.B.M. ; Liu, C.M. ; Bhojwani, S.S. - \ 2006
Plant Cell, Tissue and Organ Culture: an international journal on in vitro culture of higher plants 86 (2006)2. - ISSN 0167-6857 - p. 201 - 210.
doubled haploid plants - immediate colchicine treatment - napus cv topas - oilseed rape - desiccation tolerance - efficient production - anther cultures - heat-shock - embryos - induction
An efficient protocol to improve microspore embryogenesis is established in an important oleiferous crop, Brassica juncea (Indian mustard). Colchicine was used for enhancing microspore embryogenesis and also to obtain doubled haploid embryos. Colchicine at high concentrations (>10 mg l¿1), for 24 h, proved convenient for direct recovery of diploid embryos. Higher temperature treatment and an antiauxin PCIB (p-chlorophenoxyisobutyric acid) enhanced microspore embryogenesis significantly as compared to colchicine. An increase in temperature from 32°C to 35°C proved very efficient in increasing embryogenesis by 10-fold. The highest embryogenesis rate was obtained when PCIB was added at 35°C in the culture after 1 day of culture initiation. 20 ¿M PCIB could enhance microspore embryogenesis by 5-fold. Different abnormal shapes of embryos like lemon, banana, flask and fused cotyledons were observed. Both normal and fused cotyledonous embryos showed normal germination when transferred on the B5 basal medium
The role of sigmaB in the stress response of Gram-positive bacteria - targets for food preservation and safety
Schaik, W. van; Abee, T. - \ 2005
Current Opinion in Biotechnology 16 (2005)2. - ISSN 0958-1669 - p. 218 - 224.
listeria-monocytogenes sigma(b) - staphylococcus-aureus - bacillus-subtilis - transcription factor - biofilm formation - functional rsbu - low-temperature - acid tolerance - sigb operon - heat-shock
The alternative sigma factor ¿B modulates the stress response of several Gram-positive bacteria, including Bacillus subtilis and the food-borne human pathogens Bacillus cereus, Listeria monocytogenes and Staphylococcus aureus. In all these bacteria, ¿B is responsible for the transcription of genes that can confer stress resistance to the vegetative cell. Recent findings indicate that ¿B also plays an important role in antibiotic resistance, pathogenesis and cellular differentiation processes such as biofilm formation and sporulation. Although there are important differences in the regulation of ¿B and in the set of genes regulated by ¿B in B. subtilis, B. cereus, L. monocytogenes and S. aureus, there are also some conserved themes. A mechanistic understanding of the ¿B activation processes and assessment of its regulon could provide tools for pathogen control and inactivation both in the food industry and clinical settings
Molecular cloning and characterization of Cpn60 in the free-living nematode Plectus acuminatus
Sturzenbaum, S.R. ; Arts, M.S.J. ; Kammenga, J.E. - \ 2005
Cell Stress and Chaperones 10 (2005)2. - ISSN 1355-8145 - p. 79 - 85.
shock-protein hsp60 - heat-shock - stress-proteins - expression - gene - identification - chaperonin - atherosclerosis - autoimmune - regions
Heat shock proteins (Hsps) have provoked interest not only because of their involvement in human diseases but also for their potential as biomarkers of environmental pollution. Whereas the former interest is covered by numerous reports, the latter is an exciting new field of research. We report the isolation of the full-length cpn60 messenger ribonucleic acid (mRNA) and partial genomic deoxyribonucleic acid from the free-living, environmental sentinel nematode Plectus acuminatus, a species used in classical ecotoxicity tests. Although the primary sequence displays high identity scores to other nematodes and human Cpn60 (75% and 70%, respectively), the intron-exon structure differs markedly. Furthermore, although mRNA levels remained constant after exposure to ZnCl2 (0¿330 ¿M) under laboratory conditions, protein levels increased significantly in a dose-dependent manner. In conclusion, this first account of molecular genetic similarities and differences of Cpn60 in a neglected nematode taxon provides a valuable insight into its potential uses in gene-based ecotoxicological risk assessment exercises
Perspective: Evolution and detection of genetic robustness
Visser, J.A.G.M. de; Hermisson, J. ; Wagner, G.P. ; Ancel Meyers, L. ; Bagheri-Chaichian, H. ; Blanchard, J.L. ; Chao, L. ; Cheverud, J.M. ; Elena, S.F. ; Fontana, W. ; Gibson, G. ; Hansen, T.F. ; Krakauer, D. ; Lewontin, R.C. ; Ofria, C. ; Rice, S.H. ; Dassow, G. von; Wagner, A. ; Whitlock, M.C. - \ 2003
Evolution 57 (2003)9. - ISSN 0014-3820 - p. 1959 - 1972.
drosophila-melanogaster - developmental stability - deleterious mutations - fitness components - heat-shock - canalization - dominance - epistasis - selection - plasticity
Robustness is the invariance of phenotypes in the face of perturbation. The robustness of phenotypes appears at various levels of biological organization, including gene expression, protein folding, metabolic flux, physiological homeostasis, development, and even organismal fitness. The mechanisms underlying robustness are diverse, ranging from thermodynamic stability at the RNA and protein level to behavior at the organismal level. Phenotypes can be robust either against heritable perturbations (e.g., mutations) or nonheritable perturbations (e.g., the weather). Here we primarily focus on the first kind of robustness-genetic robustness-and survey three growing avenues of research: (1) measuring genetic robustness in nature and in the laboratory; (2) understanding the evolution of genetic robustness; and (3) exploring the implications of genetic robustness for future evolution.