B. Cornelissen | University of Amsterdam (original) (raw)

Papers by B. Cornelissen

The 12 locus in tomato confers resistance to race 2 of the soil-borne fungus Fusarium oxysporum f... more The 12 locus in tomato confers resistance to race 2 of the soil-borne fungus Fusarium oxysporum f sp lycopersici. The selective restriction fragment amplification (AFLP) positional cloning strategy was used to identify 12 in the tomato ge? nome. A yeast artificial chromosome (YAC) clone covering ^750 kb encompassing the 12 locus was isolated, and the AFLP technique was used to derive tightly linked AFLP markers from this YAC clone. Genetic complementation analy? sis in transgenic R1 plants using a set of overlapping cosmids covering the 12 locus revealed three cosmids giving full resistance to F. o. lycopersici race 2. These cosmids shared a 7-kb DNA fragment containing an open reading frame en? coding a protein with similarity to the nucleotide binding site leucine-rich repeat family of resistance genes. At the 12 locus, we identified six additional homologs that included the recently identified I2C-1 and I2C-2 genes. However, cosmids containing the I2C-1 or I2C-2 gene could not co...

Fungal Genetics and Biology, 1999

Sugar analysis of isolated cell walls from three formae speciales of Fusarium oxysporum showed th... more Sugar analysis of isolated cell walls from three formae speciales of Fusarium oxysporum showed that they not only contained glucose and (N-acetyl)-glucosamine, but also mannose, galactose and uronic acids, presumably originating from cell wall glycoproteins. Cell wall glycoproteins accounted for 50-60% of the total mass of the wall. X-ray diffraction studies showed the presence of a-l,3-glucan in the alkali-soluble cell wall fraction and of ß-1,3glucan and chitin in the alkali-insoluble fraction. Electron microscopy and lectin binding studies indicated that glycoproteins form an external layer covering an inner layer composed of chitin and glucan.

Plant Journal, 2010

Race-specific disease resistance in plants depends on the presence of resistance (R) genes. Most ... more Race-specific disease resistance in plants depends on the presence of resistance (R) genes. Most R genes encode NB-ARC-LRR proteins that carry a C-terminal leucine-rich repeat (LRR). Of the few proteins found to interact with the LRR domain, most have proposed (co)chaperone activity. Here, we report the identification of RSI2 (Required for Stability of I-2) as a protein that interacts with the LRR domain of the tomato R protein I-2. RSI2 belongs to the family of small heat shock proteins (sHSPs or HSP20s). HSP20s are ATP-independent chaperones that form oligomeric complexes with client proteins to prevent unfolding and subsequent aggregation. Silencing of RSI2-related HSP20s in Nicotiana benthamiana compromised the hypersensitive response that is normally induced by auto-active variants of I-2 and Mi-1, a second tomato R protein. As many HSP20s have chaperone properties, the involvement of RSI2 and other R protein (co)chaperones in I-2 and Mi-1 protein stability was examined. RSI2 silencing compromised the accumulation of full-length I-2 in planta, but did not affect Mi-1 levels. Silencing of heat shock protein 90 (HSP90) and SGT1 led to an almost complete loss of full-length I-2 accumulation and a reduction in Mi-1 protein levels. In contrast to SGT1 and HSP90, RSI2 silencing led to accumulation of I-2 breakdown products. This difference suggests that RSI2 and HSP90/SGT1 chaperone the I-2 protein using different molecular mechanisms. We conclude that I-2 protein function requires RSI2, either through direct interaction with, and stabilization of I-2 protein or by affecting signalling components involved in initiation of the hypersensitive response.

Plant Physiology, 2006

Resistance (R) proteins in plants confer specificity to the innate immune system. Most R proteins... more Resistance (R) proteins in plants confer specificity to the innate immune system. Most R proteins have a centrally located NB-ARC (nucleotide-binding adaptor shared by APAF-1, R proteins, and CED-4) domain. For two tomato (Lycopersicon esculentum) R proteins, I-2 and Mi-1, we have previously shown that this domain acts as an ATPase module that can hydrolyze ATP in vitro. To investigate the role of nucleotide binding and hydrolysis for the function of I-2 in planta, specific mutations were introduced in conserved motifs of the NB-ARC domain. Two mutations resulted in autoactivating proteins that induce a pathogen-independent hypersensitive response upon expression in planta. These mutant forms of I-2 were found to be impaired in ATP hydrolysis, but not in ATP binding, suggesting that the ATP- rather than the ADP-bound state of I-2 is the active form that triggers defense signaling. In addition, upon ADP binding, the protein displayed an increased affinity for ADP suggestive of a chan...

Philosophical Transactions of the Royal Society B: Biological Sciences, 1993

Virus and fungal resistance traits are important targets in the genetic engineering of agricultur... more Virus and fungal resistance traits are important targets in the genetic engineering of agricultural and horticultural crops. We have engineered resistance against potato virus X in important commercial potato cultivars. Four years of field trials with resistant potatoes have demonstrated the commercial feasibility of improving potato cultivars by selectively adding new traits while preserving intrinsic properties. In our pursuit for a broad resistance against fungi we have focused on the exploitation of genes encoding antifungal proteins. We present results demonstrating the antifungal effect of some of these proteins in vitro , as well as the synergy between specific chitinases and β-1,3-glucanases. We also report high level resistance against Fusarium oxysporum in transgenic tomato plants expressing a specific combination of genes encoding these enzymes.

Euphytica, 1995

UvA-DARE (Digital Academic Repository) Synergistic activity of chitinases and beta-1,3-glucanases... more UvA-DARE (Digital Academic Repository) Synergistic activity of chitinases and beta-1,3-glucanases enhances fungal resistance in transgenic tomato plants

Plant Molecular Biology, 1993

The Nicotiana tabacum ap24 gene encoding a protein with antifungal activity toward Phytophthora i... more The Nicotiana tabacum ap24 gene encoding a protein with antifungal activity toward Phytophthora infestans has been characterized. Analysis of cDNA clones revealed that at least three ap24-like genes are induced in tobacco upon infection with tobacco mosaic virus. Amino acid sequencing of the purified protein showed that AP24 is synthesized as a preproprotein from which an amino-terminal signal peptide and a carboxyl-terminal propeptide (CTPP) are cleaved off during post-translational processing. The functional role of the CTPP was investigated by expressing chimeric genes encoding either wild-type AP24 or a mutant protein lacking the CTPP. Plants expressing the wild-type construct resulted in proteins properly sorted to the vacuole. In contrast, the proteins produced in plants expressing the mutant construct were secreted extracellularly, indicating that the CTPP is necessary for targeting of AP24 to the vacuoles. Similar results were obtained for vacuolar chitinases and beta-1,3-glucanases of tobacco. The extracellularly targeted mutant proteins were shown to have retained their biological activity. Together, these results suggest that within all vacuolar pathogenesis-related proteins the targeting information resides in a short carboxyl-terminal propeptide which is removed during or after transport to the plant vacuole.

A nove1 pathogen- and wound-inducible antifungal protein of 20 kD was purified from tobacco (Nico... more A nove1 pathogen- and wound-inducible antifungal protein of 20 kD was purified from tobacco (Nicotiana tabacum) Samsun NN leaves inoculated with tobacco mosaic virus (TMV). The protein, designated CBPZO, was purified by chitin-affinity chromatography and gel filtration. In vitro assays demonstrated that CBPZO exhibits antifungal activity toward Trichoderma viride and Fusarium solani by causing lysis of the germ tubes and/or

Plant Molecular Biology 2, 1991

Extensive potato breeding programs over the years have yielded improved varieties with a whole se... more Extensive potato breeding programs over the years have yielded improved varieties with a whole set of proven valuable traits. The process of potato breeding, however, is laborious and time-consuming due to the tetraploid character of the potato genome. Techniques for the genetic engineering of plants present a new tool to improve potato via the introduction of traits presently missing in the existing cultivars. An essential element in the successful application of these new techniques is the preservation of the existing, desired traits of the cultivars. So far, no systematic studies on the actual performance of engineered plants have been performed. In this paper we report the engineering of resistance to potato virus X in the two commercial potato cultivars Bintje and Escort. Also, the results of two years of field trials testing the performance of these transgenic lines are discussed.

New Journal of Chemistry - NEW J CHEM, 1997

Plant Physiology, 1999

The gene encoding the precursor to stinging nettle (Urtica dioica L.) isolectin I was introduced ... more The gene encoding the precursor to stinging nettle (Urtica dioica L.) isolectin I was introduced into tobacco (Nicotiana tabacum). In transgenic plants this precursor was processed to mature-sized lectin. The mature isolectin is deposited intracellularly, most likely in the vacuoles. A gene construct lacking the C-terminal 25 amino acids was also introduced in tobacco to study the role of the C terminus in subcellular trafficking. In tobacco plants that expressed this construct, the mutant precursor was correctly processed and the mature isolectin was targeted to the intercellular space. These results indicate the presence of a C-terminal signal for intracellular retention of stinging nettle lectin and most likely for sorting of the lectin to the vacuoles. In addition, correct processing of this lectin did not depend on vacuolar deposition. Isolectin I purified from tobacco displayed identical biological activities as isolectin I isolated from stinging nettle. In vitro antifungal as...

Molecular Genetics and Genomics, 2002

In order to genetically map and eventually isolate avirulence genes, parasexual crosses between d... more In order to genetically map and eventually isolate avirulence genes, parasexual crosses between different races of Fusarium oxysporum f. sp. lycopersici were performed by means of protoplast fusion. Two wild-type strains, race 1 Fol004 (A1a2a3) and race 3 Fol029 (a1a2A3), were transformed with phleomycin and hygromycin resistance genes, respectively. In total 32 fusion products were selected by screening for the presence of both marker genes. The presence of either avirulence gene A1 or A3 in the fusion products was determined by plant bioassays. Segregation of avirulence revealed a bias for the presence of A1. Two recombinants for the avirulence phenotype were observed, each with a new association of avirulence genes never observed to exist in the wild. Electrophoretic karyotype analysis revealed that chromosome patterns were different for all fusion products. Hybridization patterns using various probes indicated that chromosome rearrangements and recombination had occurred. Karyotype analysis of the two avirulence recombinants revealed hybrid karyotypes resulting from a massive exchange of parental DNA. This indicates that the present population of recombinants can be used for gene mapping in the asexual fungus F. oxysporum f. sp. lycopersici.

FEBS Letters, 2002

The coding sequence of a major xylem sap protein of tomato was identi¢ed with the aid of mass spe... more The coding sequence of a major xylem sap protein of tomato was identi¢ed with the aid of mass spectrometry. The protein, XSP10, represents a novel family of extracellular plant proteins with structural similarity to plant lipid transfer proteins. The XSP10 gene is constitutively expressed in roots and lower stems. The decline of XSP10 protein levels in tomato infected with a fungal vascular pathogen may re£ect breakdown or modi¢cation by the pathogen.

Environmental Microbiology, 2008

Fusarium oxysporum is an asexual fungus that inhabits soils throughout the world. As a species, F... more Fusarium oxysporum is an asexual fungus that inhabits soils throughout the world. As a species, F. oxysporum can infect a very broad range of plants and cause wilt or root rot disease. Single isolates of F. oxysporum, however, usually infect one or a few plant species only. They have therefore been grouped into formae speciales (f.sp.) based on host specificity. Isolates able to cause tomato wilt (f.sp. lycopersici) do not have a single common ancestor within the F. oxysporum species complex. Here we show that, despite their polyphyletic origin, isolates belonging to f.sp. lycopersici all contain an identical genomic region of at least 8 kb that is absent in other formae speciales and non-pathogenic isolates, and comprises the genes SIX1, SIX2 and SHH1. In addition, SIX3, which lies elsewhere on the same chromosome, is also unique for f.sp. lycopersici. SIX1 encodes a virulence factor towards tomato, and the Six1, Six2 and Six3 proteins are secreted in xylem during colonization of tomato plants. We speculate that these genes may be part of a larger, dispensable region of the genome that confers the ability to cause tomato wilt and has spread among clonal lines of F. oxysporum through horizontal gene transfer. Our findings also have practical implications for the detection and identification of f.sp. lycopersici.

Molecular Genetics and Genomics, 2001

As part of an investigation of the cell wall structure of plant pathogenic, filamentous fungi, we... more As part of an investigation of the cell wall structure of plant pathogenic, filamentous fungi, we set out to characterize covalently bound cell wall glycoproteins (CWPs) of the tomato pathogen Fusarium oxysporum. N-terminal sequencing of an abundant 60-kDa CWP led to the cloning of the corresponding gene, which we have designated FEM1 (Fusarium extracellular matrix protein). The gene contains an ORF encoding a primary translation product of 212 amino acids, including an N-terminal 17-amino acid secretion signal sequence. Furthermore, FEM1p contains two potential N-glycosylation sites, and is rich in serine and threonine residues (29%) that could serve as O-glycosyl addition sites. At its C-terminus the protein contains a 22-amino acid sequence with the characteristics of a glycosyl-phosphatidylinositol (GPI) anchor addition signal. A mutant FEM1 protein lacking this GPI anchor addition signal is not retained in the fungal cell wall but released into the culture medium, indicating that in the wild-type protein this sequence functions to anchor the protein to the extracellular matrix. Southern analysis shows that FEM1 is present as a single-copy gene in all formae speciales of F. oxysporum tested and in F. solani. Database searches show that FEM1p homologous sequences are present in other filamentous fungi as well.