Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

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    We will mail you new results for this query: keywords==gene-transfer
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TCR's genetically linked to CD28 and CD3e do not mispair with endogous TCR chains and mediate enhanced T cell persistance and anti-melanoma activity
Govers, C.C.F.M. ; Sebestyen, Z. ; Roszik, J. ; Brakel, M. van; Berrevoets, C. ; Szoor, A. ; Panoutsopoulou, K. ; Broertjes, M. ; Van, T. ; Vereb, G. ; Szollosi, J. ; Debets, R. - \ 2014
The Journal of Immunology 193 (2014)10. - ISSN 0022-1767 - p. 5315 - 5326.
chimeric-antigen-receptor - gene-transfer - metastatic melanoma - cancer regression - lymphocytes - therapy - alpha - activation - toxicity - survival
Adoptive transfer of T cells that are gene engineered to express a defined TCR represents a feasible and promising therapy for patients with tumors. However, TCR gene therapy is hindered by the transient presence and effectiveness of transferred T cells, which are anticipated to be improved by adequate T cell costimulation. In this article, we report the identification and characterization of a novel two-chain TCR linked to CD28 and CD3e (i.e., TCR:28e). This modified TCR demonstrates enhanced binding of peptide–MHC and mediates enhanced T cell function following stimulation with peptide compared with wild-type TCR. Surface expression of TCR:28e depends on the transmembrane domain of CD28, whereas T cell functions depend on the intracellular domains of both CD28 and CD3e, with IL-2 production showing dependency on CD28:LCK binding. TCR:28e, but not wild-type TCR, induces detectable immune synapses in primary human T cells, and such immune synapses show significantly enhanced accumulation of TCR transgenes and markers of early TCR signaling, such as phosphorylated LCK and ERK. Importantly, TCR:28e does not show signs of off-target recognition, as evidenced by lack of TCR mispairing, as well as preserved specificity. Notably, when testing TCR:28e in immune-competent mice, we observed a drastic increase in T cell survival, which was accompanied by regression of large melanomas with limited recurrence. Our data argue that TCR transgenes that contain CD28, and, thereby, may provide T cell costimulation in an immune-suppressive environment, represent candidate receptors to treat patients with tumors.
Efficiency of Agrobacterium rhizogenes-mediated root transformation of Parasponia and Trema is temperature dependent
Cao, Q. ; Camp, R. Op den; Seifi Kalhor, M. ; Bisseling, T. ; Geurts, R. - \ 2012
Plant Growth Regulation 68 (2012)3. - ISSN 0167-6903 - p. 459 - 465.
medicago-truncatula - gene-transfer - non-legume - plants - nitrogen - andersonii - nodulation - rhizobium - phaseolus - sequences
Parasponia trees are the only non-legume species that form nitrogen-fixing root nodules with rhizobium. Based on its taxonomic position in relation to legumes (Fabaceae), it is most likely that both lineages have gained this symbiotic capacity independently. Therefore, Parasponia forms a bridging species to understand the evolutionary constraints underlying this symbiosis. However, absence of key technologies to genetically modify Parasponia seriously impeded studies on these species. We employed Agrobacterium rhizogenes to create composite Parasponia andersonii plants that harbour transgenic roots. Here, we provide an optimized protocol to infect P. andersonii as well as its non-symbiotic sister species Trema tomentosa with A. rhizogenes. We show that the transformation efficiency is temperature dependent. Whereas the optimal growth temperature for both these species is 28 °C, the transformation is most efficient when co-cultivation with A. rhizogenes occurs at 21 °C. Using this optimized protocol up to 80 % transformation efficiency can be obtained. These robust transformation platforms will provide a strong tool to unravel the Parasponia–rhizobium symbiosis
Effector diversification within compartments of the Leptosphaeria maculans genome affected by repeat induced point mutations
Rouxel, T. ; Grandaubert, J. ; Hane, J.K. ; Hoede, C. ; Wouw, A. ; Couloux, A. ; Dominguez, V. ; Anthouard, V. ; Bally, P. ; Bourras, S. ; Cozijnsen, A.J. ; Ciuffetti, L.M. ; Degrave, A. ; Dilmaghani, A. ; Duret, L. ; Fudal, L. ; Goodwin, S.B. ; Gout, L. ; Glaser, N. ; Linglin, J. ; Kema, G.H.J. ; Lapalu, N. ; Lawrence, C.B. ; May, K. ; Meyer, M. ; Ollivier, B. ; Poulain, J. ; Schoch, C.L. ; Simon, A. ; Spatafora, J.W. ; Stachowiak, A. ; Turgeon, B.G. ; Tyler, B.M. ; Vincent, D. ; Weissenbach, J. ; Amselem, J. ; Quesneville, H. ; Oliver, R.P. ; Wincker, P. ; Balesdent, M.H. ; Howlett, B.J. - \ 2011
Nature Communications 2 (2011). - ISSN 2041-1723 - p. 202 - 202.
transposable elements - molecular evolution - pathogen effectors - brassica-napus - gene-transfer - oilseed rape - stem canker - avirulence - plant - fungal
Fungi are of primary ecological, biotechnological and economic importance. Many fundamental biological processes that are shared by animals and fungi are studied in fungi due to their experimental tractability. Many fungi are pathogens or mutualists and are model systems to analyse effector genes and their mechanisms of diversification. In this study, we report the genome sequence of the phytopathogenic ascomycete Leptosphaeria maculans and characterize its repertoire of protein effectors. The L. maculans genome has an unusual bipartite structure with alternating distinct guanine and cytosine-equilibrated and adenine and thymine (AT)-rich blocks of homogenous nucleotide composition. The AT-rich blocks comprise one-third of the genome and contain effector genes and families of transposable elements, both of which are affected by repeat-induced point mutation, a fungal-specific genome defence mechanism. This genomic environment for effectors promotes rapid sequence diversification and underpins the evolutionary potential of the fungus to adapt rapidly to novel host-derived constraints
A novel mode of chromosomal evolution peculiar to filamentous Ascomycete fungi
Hane, J.K. ; Rouxel, T. ; Howlett, B.J. ; Kema, G.H.J. ; Goodwin, S.B. ; Oliver, R.P. - \ 2011
Genome Biology 12 (2011). - ISSN 1474-7596 - 16 p.
pathogen stagonospora-nodorum - genome sequence - medicago-truncatula - neurospora-crassa - gene-transfer - wheat - phylogeny - synteny - conservation - organization
Background - Gene loss, inversions, translocations, and other chromosomal rearrangements vary among species, resulting in different rates of structural genome evolution. Major chromosomal rearrangements are rare in most eukaryotes, giving large regions with the same genes in the same order and orientation across species. These regions of macrosynteny have been very useful for locating homologous genes in different species and to guide the assembly of genome sequences. Previous analyses in the fungi have indicated that macrosynteny is rare; instead, comparisons across species show no synteny or only microsyntenic regions encompassing usually five or fewer genes. To test the hypothesis that chromosomal evolution is different in the fungi compared to other eukaryotes, synteny was compared between species of the major fungal taxa. Results - These analyses identified a novel form of evolution in which genes are conserved within homologous chromosomes, but with randomized orders and orientations. This mode of evolution is designated mesosynteny, to differentiate it from micro- and macrosynteny seen in other organisms. Mesosynteny is an alternative evolutionary pathway very different from macrosyntenic conservation. Surprisingly, mesosynteny was not found in all fungal groups. Instead, mesosynteny appears to be restricted to filamentous Ascomycetes and was most striking between species in the Dothideomycetes. Conclusions - The existence of mesosynteny between relatively distantly related Ascomycetes could be explained by a high frequency of chromosomal inversions, but translocations must be extremely rare. The mechanism for this phenomenon is not known, but presumably involves generation of frequent inversions during meiosis
Bias and conflict in phylogenetic inference of myco-heterotrophic plants: a case study in Thismiaceae
Merckx, V. ; Bakker, F.T. ; Huysmans, K. ; Smets, B.F. - \ 2009
Cladistics-The International Journal of the Willi Hennig Society 25 (2009)1. - ISSN 0748-3007 - p. 64 - 77.
long-branch attraction - 18s rdna sequences - parasitic plants - nucleotide substitution - flowering plants - tree selection - molecular-data - gene-transfer - data sets - burmanniaceae
Due to morphological reduction and absence of amplifiable plastid genes, the identification of photosynthetic relatives of heterotrophic plants is problematic. Although nuclear and mitochondrial gene sequences may offer a welcome alternative source of phylogenetic markers, the presence of rate heterogeneity in these genes may introduce bias/systematic error in phylogenetic analyses. We examine the phylogenetic position of Thismiaceae based on nuclear 18S rDNA and mitochondrial atpA DNA sequence data, as well as using parsimony, likelihood and Bayesian inference methods. Significant differences in evolutionary rates of these genes between closely related taxa lead to conflicting results: while parsimony analyses of 18S rDNA and combined data strongly support the monophyly of Thismiaceae, Bayesian inference, with and without a relaxed molecular clock, as well as the Swofford-Olsen-Waddell-Hillis (SOWH) test confidently reject this hypothesis. We show that rate heterogeneity in our data leads to long-branch attraction artifacts in parsimony analysis. However, using model-based inference methods the question of whether Thismiaceae are monophyletic remains elusive. On the one hand maximum likelihood nonparametric bootstrapping and parametric hypothesis tests fail to support a paraphyletic Thismiaceae, on the other hand Bayesian inference methods (both without and with a relaxed clock) significantly reject a monophyletic Thismiaceae. These results show that an adequate sampling, the use of rate homogeneous data, and the application of different inference methods are important factors for developing phylogenetic hypotheses of myco-heterotrophic plants
The pOT and pLOB vector systems: Improving ease of transgene expression in Botrytis cinerea
Patel, R.M. ; Heneghan, M.N. ; Kan, J.A.L. van; Bailey, A.M. ; Foster, G.D. - \ 2008
Journal of General and Applied Microbiology 54 (2008)6. - ISSN 0022-1260 - p. 367 - 376.
green-fluorescent protein - agrobacterium-mediated transformation - heat-shock response - coprinus-cinereus - gene-transfer - reporter - gfp - mutations - plants - resistance
This paper outlines the construction of a novel vector system comprising interchangeable terminators, as well as a multiple cloning site (MCS), to facilitate the transformation of the fungal plant pathogen Botrytis cinerea. Previous molecular studies on B. cinerea have relied upon the pLOB1 based vector system (controlled by the Aspergillus nidulans oliC promoter and a region reported to be the B. cinerea tubA terminator). Investigations, however, have revealed that, rather than the genuine B. cinerea tubA terminator, the pLOB1 terminator fragment is from another gene locus within the genome. Because previous studies have found that terminators aide in transcript stability, the main aims of this study were to develop and evaluate both vector systems, pOT (controlled by the A. nidulans oliC promoter and A. nidulans trpC terminator) and pLOB, with a range of exogenous genes, including enhanced green fluorescent protein (eGFP), monomeric red fluorescent protein (mRFP), luciferase (LUC) and ß-glucuronidase (GUS). Our investigations demonstrate that pLOB and pOT based vectors are capable of expressing all four reporter genes and may be applied to future molecular studies on B. cinerea and other related ascomycetes. Additionally, this is the first reported expression of mRFP and LUC in B. cinerea.
Angiopoietin-like-4 is a potential angiogenic mediator in arthritis
Hermann, L.M. ; Pinkerton, M. ; Jennings, K. ; Yang, L. ; Grom, A. ; Sowders, D. ; Kersten, A.H. ; Witte, D.P. ; Hirsch, R. ; Thornton, S. - \ 2005
Clinical Immunology 115 (2005)1 sp. is.. - ISSN 1521-6616 - p. 93 - 101.
collagen-induced arthritis - endothelial growth-factor - necrosis-factor-alpha - rheumatoid-arthritis - gene-transfer - target gene - expression - inhibition - protein - vegf
Our previous studies of gene expression profiling during collagen-induced arthritis (CIA) indicated that the putative angiogenic factor Angptl4 was one of the most highly expressed mRNAs early in disease. To investigate the potential involvement of Angptl4 in CIA pathogenesis, Angptl4 protein levels were assessed at early stages of disease and its cellular sources were determined. In addition, the functional effects of mouse Angptl4 on endothelial cells were assessed. Angptl4 protein levels were higher in arthritic joints as compared to normal joints. In situ hybridization localized Angptl4 mRNA to stromal fibroblast-like cells within the inflamed synovium. Temporal expression of Angptl4 mRNA during CIA was similar to that of key angiogenic factors, including structurally related angiopoietin 1. Recombinant mouse Angptl4 promoted endothelial cell survival and formation of tubule-like structures. These functional effects of Angptl4, combined with very high expression at early stages of CIA, suggest a role for Angptl4 in angiogenesis in arthritis
Cre recombinase expression can result in phenotypic aberrations in plants
Coppoolse, E. ; Vroomen, M.J. de; Roelofs, D. ; Smit, J. ; Gennip, F. van; Hersmus, B.J.M. ; Nijkamp, H.J.J. ; Haaren, M.J. van - \ 2003
Plant Molecular Biology 51 (2003)2. - ISSN 0167-4412 - p. 263 - 279.
site-specific recombination - agrobacterium-tumefaciens - transgenic tobacco - gene-transfer - dna - genome - transformation - selection - vectors - lox
The cre recombinase gene was stably introduced and expressed in tomato, petunia and Nicotiana tabacum. Some plants expressing the cre gene driven by a CaMV 35S promoter displayed growth retardation and a distinct pattern of chlorosis in their leaves. Although no direct relation can be proven between the phenotype and cre expression, aberrant phenotypes always co-segregate with the transgene, which strongly suggests a correlation. The severity of the phenotype does not correlate with the level of steady-state mRNA in mature leaves, but with the timing of cre expression during organogenesis. The early onset of cre expression in tomato is correlated with a more severe phenotype and with higher germinal transmission frequencies of site-specific deletions. No aberrant phenotype was observed when a tissue-specific phaseolin promoter was used to drive the cre gene. The data suggest that for the application of recombinases in plants, expression is best limited to specific tissues and a short time frame.[12pt] Abbreviations: bar, the phosphinotricin acetyltransferase gene; CAM, chloramphenicol resistance gene; Ds 5 & Ds 3, borders of the Ds transposable element from maize forming a functional transposable element that embodies the interjacent DNA; gus, the -glucoronidase gene; gus-int, the gus gene interrupted by a plant intron; hpt, the hygromycin phosphotransferase gene; nptII, the neomycin phosphotransferase gene; ORI, bacterial origin for plasmid replication in Escherichia coli of plasmid p15A Cre recombinase - lox P - petunia - site-specific recombination - tobacco - tomato - toxicity
Agrobacterium tumefaciens mediated transformation of Allium cepa L.: the production of transgenic onions and shallots
Zheng, S.J. ; Khrustaleva, L.I. ; Henken, G. ; Sofiari, E. ; Jacobsen, E. ; Kik, C. ; Krens, F.A. - \ 2001
Molecular Breeding 7 (2001)2. - ISSN 1380-3743 - p. 101 - 115.
manihot-esculenta crantz - oryza-sativa-l - gene-transfer - indica rice - t-dna - transient expression - suspension-cultures - plant-regeneration - immature embryos - reporter gene
This paper describes the development of a reliable transformation protocol for onion and shallot (Allium cepa L.) which can be used year-round. It is based on Agrobacterium tumefaciens as a vector, with three-week old callus, induced from mature zygotic embryos, as target tissue. For the development of the protocol a large number of parameters were studied. The expression of the uidA gene coding for -glucuronidase was used as an indicator in the optimization of the protocol. Subspecies (onion and shallot) and cultivar were important factors for a successful transformation: shallot was better than onion and for shallot cv. Kuning the best results were obtained. Also, it was found that constantly reducing the size of the calli during subculturing and selection by chopping, thus enhancing exposure to the selective agent hygromycin, improved the selection efficiency significantly. Furthermore, callus induction medium and co-cultivation period showed a significant effect on successful stable transformation. The usage of different Agrobacterium strains, callus ages, callus sources and osmotic treatments during co-cultivation did not influence transformation efficiency. The highest transformation frequency (1.95%), was obtained with shallot cv. Kuning. A total of 11 independent transformed callus lines derived from zygotic embryos were produced: seven lines from shallot and four lines from onion. Large differences in plantlet production were observed among these lines. The best line produced over 90 plantlets. Via PCR the presence of the uidA and hpt (hygromycin phosphotransferase) genes could be demonstrated in these putative transformed plants. Southern hybridization showed that most lines originated from one transformation event. However, in one line plants were obtained indicating the occurrence and rescue of at least three independent transformation events. This suggested that T-DNA integration occurred in different cells within the callus. Most transgenic plants only had one copy of T-DNA integrated into their genomes. FISH performed on 12 plants from two different lines representing two integration events showed that original T-DNA integration had taken place on the distal end of chromosomes 1 or 5. A total of 83 transgenic plants were transferred to the greenhouse and these plants appeared to be diploid and normal in morphology
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