[How do disease resistance genes function?].
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Genes, Plant↗
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[The structure and regulation of plant proteinase inhibitor genes and their strategy in pest control].
The diverse plant proteinase inhibitors (PIs) genes from different plant species have been isolated and their products with one or more genes are targeted at different biochemical and physiological process with the insect, and well played a potent defensive role against insects and pathogens. The use of recombinant PIs and synergistic activation to protect plants has been incorporated in integrated pest management program. Though they may not replace the use of chemical pesticides in the near future, but effectively complement it. Currently, the role and mechanism of action for most of these inhibitors are being studied in detail. This article describes the structure of PIs genes, their regulation and expression, and discusses the strategy to develop transgenic plants against phytophagous insects.
Animals↗
Clinostating effects on Apogee wheat resistance to wheat streak mosaic virus.
The effects of vertical and horizontal clinostating with container velocity 2 rpm and platform velocity 1 rpm on Apogee wheat resistance to wheat streak mosaic virus (WSMV) were studied. For the first time the yield of grain was obtained from both, healthy and the WSMV-infected wheat plants under conditions of simulated microgravity (clinorotation) which is accounted for by the inclusion of various mechanisms in the process of plant adaptation to the factors of stresses. For the first time the stages of viral infection development were elucidated under the conditions of prolonged clinostating and the presence of the WSMV was detected in wheat plants on the 18th day after inoculation employing the method of polymerase chain reaction (PCR). In the test variant with vertical clinostating (R=1,0) the least favorable conditions for viral reproduction were identified. In the same variant the increases of the total nitrogen content in the ears were observed which may be an evidence of interferon-like protein synthesis in plant similar to the interferon system functioning in animals.
Biomass↗
Altered immune status in aluminum reduction plant workers.
From 1978 to 1985, B-cell lymphoma occurred in five employees of an aluminum reduction plant (expected = 0.2; Poisson distribution). As immunodeficiency is a known risk factor for B-cell lymphoma, we did a pilot study to evaluate immune function in apparently healthy plant workers. Twenty-three volunteers were selected for study from 350 workers, representing a range of experience in the potroom and with exposures to strong magnetic fields and volatilized aromatic hydrocarbons. Potroom workers had significantly higher T8 levels (mean = 1,227) than non-potroom workers (mean = 597) (p less than .05, Wilcoxon rank sums) or established normal values (median = 450). T4 levels were higher for potroom workers (mean = 1,017) than for non-potroom workers (mean = 558) or for established norms (median = 756) (p less than .10, Wilcoxon rank sums). Ten of 20 potroom workers had abnormal T4/T8 ratios (less than 0.91) due to disproportionate elevation of the T8 subpopulation. These data suggest an underlying immune alteration in the aluminum workers studied. Further study is needed to assess the implications of abnormal T-cell subsets in a worker population with high rates of lymphoma.
Adult↗
Immunization against rabies with plant-derived antigen.
We previously demonstrated that recombinant plant virus particles containing a chimeric peptide representing two rabies virus epitopes stimulate virus neutralizing antibody synthesis in immunized mice. We show here that mice immunized intraperitoneally or orally (by gastric intubation or by feeding on virus-infected spinach leaves) with engineered plant virus particles containing rabies antigen mount a local and systemic immune response. After the third dose of antigen, given intraperitoneally, 40% of the mice were protected against challenge infection with a lethal dose of rabies virus. Oral administration of the antigen stimulated serum IgG and IgA synthesis and ameliorated the clinical signs caused by intranasal infection with an attenuated rabies virus strain.
Administration, Oral↗
Identification of components in disease-resistance signaling in Arabidopsis by map-based cloning.
With the whole genome sequence and thousands of defined polymorphisms between ecotypes available, it has become much easier to clone a gene by position (map-based cloning) in Arabidopsis. Recent development of DNA-isolation methods in plants also dramatically facilitated large-scale processing of DNA samples. Here, we describe detailed protocols for each step on general scheme of map-based cloning, from mutagenesis to genetic analysis, from rough mapping to fine mapping, and at the end to cloning the gene. Not only can these methods be used to isolate genes that are involved in plant innate immunity, they can also be adapted for any forward genetics projects in Arabidopsis.
Arabidopsis↗
Clinical role of lipid transfer proteins in food allergy.
Lipid transfer proteins are widespread plant food allergens, highly resistant to food processing and to the gastrointestinal environment, which have recently been described as true food allergens in the Mediterranean area, where they have been associated with severe allergic reactions to foods in patients without pollen allergy. In this review we analyze their molecular structure, biological function, and clinical relevance in food allergy.
Antigens, Plant↗
Genetic and physical mapping of Pi37(t), a new gene conferring resistance to rice blast in the famous cultivar St. No. 1.
The famous rice cultivar (cv.), St. No. 1, confers complete resistance to many isolates collected from the South China region. To effectively utilize the resistance, a linkage assay using microsatellite markers (SSR) was performed in the three F2 populations derived from crosses between the donor cv. St. No. 1 and each of the three susceptible cvs. C101PKT, CO39 and AS20-1, which segregated into 3R:1S (resistant/susceptible) ratio, respectively. A total of 180 SSR markers selected from each chromosome equally were screened. The result showed that the two markers RM128 and RM486 located on chromosome 1 were linked to the resistance gene in the respective populations above. This result is not consistent with those previously reported, in which a well-known resistance gene Pif in the St. No. 1 is located on chromosome 11. To confirm this result, additional four SSR markers, which located in the region lanked by RM128 and RM486, were tested. The results showed that markers RM543 and RM319 were closer to, and RM302 and RM212 completely co-segregated with the resistance locus detected in the present study. These results indicated that another resistance gene involved in the St. No. 1, which is located on chromosome 1, and therefore tentatively designated as Pi37(t). To narrow down genomic region of the Pi37(t) locus, eight markers were newly developed in the target region through bioinformatics analysis (BIA) using the publicly available sequences. The linkage analysis with these markers showed that the Pi37(t) locus was mapped to a approximately 0.8 centimorgans (cM) interval flanked by RM543 and FPSM1, where a total of seven markers co-segregated with it. To physically map the locus, the Pi37(t)-linked markers were landed on the reference sequence of cv. Nipponbare through BIA. A contig map corresponding to the locus was constructed based on the reference sequence aligned by the Pi37(t)-linked markers. Consequently, the Pi37(t) locus was defined to 374 kb interval flanking markers RM543 and FPSM1, where only four candidate genes with the resistance gene conserved structure (NBS-LRR) were further identified to a DNA fragment of 60 kb in length by BIA.
China↗
Identification of functional apple scab resistance gene promoters.
Apple scab (Venturia inaequalis) is one of the most damaging diseases affecting commercial apple production. Some wild Malus species possess resistance against apple scab. One gene, HcrVf2, from a cluster of three genes derived from the wild apple Malus floribunda clone 821, has recently been shown to confer resistance to apple scab when transferred into a scab-susceptible apple variety. For this proof-of-function experiment, the use of the 35S promoter from Cauliflower mosaic virus was reliable and appropriate. However, in order to reduce the amount of non-plant DNA in genetically modified apple to a minimum, with the aim of increasing genetically modified organism acceptability, these genes would ideally be regulated by their own promoters. In this study, sequences from the promoter region of the three members of the HcrVf gene family were compared. Promoter constructs containing progressive 5' deletions were prepared and used for functional analyses. Qualitative assessment confirmed promoter activity in apple. Quantitative promoter comparison was carried out in tobacco (Nicotiana glutinosa) and led to the identification of several promoter regions with different strengths from a basal level to half the strength of the 35S promoter from Cauliflower mosaic virus.
Ascomycota↗
Genetic control of rice blast resistance in the durably resistant cultivar Gumei 2 against multiple isolates.
To further our understanding of the genetic control of blast resistance in rice cultivar Gumei 2 and, consequently, to facilitate the utilization of this durably blast-resistant cultivar, we studied 304 recombinant inbred lines of indica rice cross Zhong 156/Gumei 2 and a linkage map comprising 181 markers. An analysis of segregation for resistance against five isolates of rice blast suggested that one gene cluster and three additional major genes that are independently inherited are responsible for the complete resistance of Gumei 2. The gene cluster was located to chromosome 6 and includes two genes mapped previously, Pi25(t), against Chinese rice blast isolate 92-183 (race ZC15) and Pi26(t) against Philippine rice blast isolate Ca89 (lineage 4), and a gene for resistance against Philippine rice blast isolate 92330-5 (lineage 17). Of the two genes conferring resistance against the Philippine isolates V86013 (lineage 15) and C923-39 (lineage 46), we identified one as Pi26(t) and mapped the other onto the distal end of chromosome 2 where Pib is located. We used three components of partial blast resistance, percentage diseased leaf area (DLA), lesion number and lesion size, all measured in the greenhouse, to measure the degree of susceptibility to isolates Ca89 and C923-39 and subsequently identified nine and eight quantitative trait loci (QTLs), respectively. Epistasis was determined to play an important role in partial resistance against Ca89. Using DLA measured on lines susceptible in a blast nursery, we detected six QTLs. While different QTLs were detected for partial resistance to Ca89 and C923-39, respectively, most were involved in the partial resistance in the field. Our results suggest that the blast resistance in Gumei 2 is controlled by multiple major genes and minor genes with epistatic effects.
Chromosome Mapping↗
Identification of a large cluster of coiled coil-nucleotide binding site--leucine rich repeat-type genes from the Rps1 region containing Phytophthora resistance genes in soybean.
Fifteen Rps genes confer resistance against the oomycete pathogen Phytophthora sojae, which causes root and stem rot disease in soybean. We have isolated a disease resistance gene-like sequence from the genomic region containing Rps1-k. Four classes of cDNA of the sequence were isolated from etiolated hypocotyl tissues that express the Rps1-k-encoded Phytophthora resistance. Sequence analyses of a cDNA clone showed that the sequence is a member of the coiled coil-nucleotide binding site-leucine rich repeat (CC-NBS-LRR)-type of disease resistance genes. It showed 36% identity to the recently cloned soybean resistance gene Rpg1-b, which confers resistance against Pseudomonas syringae pv. glycinea, and 56% and 38% sequence identity to putative resistance gene sequences from lotus and Medicago truncatula, respectively. The soybean genome contains about 38 copies of the sequence. Most of these copies are clustered in approximately 600 kb of contiguous DNA of the Rps1-k region. We have identified a recombinant that carries both rps1-k- and Rps1-k-haplotype-specific allelomorphs of two Rps1-k-linked molecular markers. An unequal crossover event presumably led to duplication of alleles for these two physically linked molecular markers. We hypothesize that the unequal crossing over was one of the mechanisms involved in tandem duplication of CC-NBS-LRR sequences in the Rps1-k region.
Amino Acid Sequence↗
Powdery mildew resistance and Lr34/Yr18 genes for durable resistance to leaf and stripe rust cosegregate at a locus on the short arm of chromosome 7D of wheat.
The incorporation of effective and durable disease resistance is an important breeding objective for wheat improvement. The leaf rust resistance gene Lr34 and stripe rust resistance gene Yr18 are effective at the adult plant stage and have provided moderate levels of durable resistance to leaf rust caused by Puccinia triticina Eriks. and to stripe rust caused by Puccinia striiformis Westend. f. sp. tritici. These genes have not been separated by recombination and map to chromosome 7DS in wheat. In a population of 110 F(7) lines derived from a Thatcher x Thatcher isogenic line with Lr34/Yr18, field resistance to leaf rust conferred by Lr34 and to stripe rust resistance conferred by Yr18 cosegregated with adult plant resistance to powdery mildew caused by Blumeria graminis (DC) EO Speer f. sp. tritici. Lr34 and Yr18 were previously shown to be associated with enhanced stem rust resistance and tolerance to barley yellow dwarf virus infection. This chromosomal region in wheat has now been linked with resistance to five different pathogens. The Lr34/Yr18 phenotypes and associated powdery mildew resistance were mapped to a single locus flanked by microsatellite loci Xgwm1220 and Xgwm295 on chromosome 7DS.
Chromosome Mapping↗
Gene discovery and microarray analysis of cacao (Theobroma cacao L.) varieties.
The cacao bean harvest from the relatively under developed tropical tree cacao (Theobroma cacao L.) is subject to high losses in potential production due to pests and diseases. To discover and understand the stability of putative natural resistance mechanisms in this commodity crop, essential for chocolate production, we undertook a gene-discovery program and demonstrated its use in gene-expression arrays. Sequencing and assembling bean and leaf cDNA library inserts produced a unique contig set of 1,380 members. High-quality annotation of this gene set using Blast and MetaFam produced annotation for 75% of the contigs and allowed us to identify the types of gene expressed in cacao beans and leaves. Microarrays were constructed using amplified inserts of the uni-gene set and challenged with bean and leaf RNA from five cacao varieties. The microarray performed well across the five randomly chosen cacao genotypes and did not show a bias towards either leaf or bean tissues. This demonstrates that the gene sequences are useful for microarray analysis across cacao genotypes and tissue types. The array results, when compared with real-time PCR results for selected genes, showed a correlation with differential gene-expression patterns. We intend that the resultant DNA sequences and molecular microarray platform will help the cacao community to understand the basis, likely stability and pathotype resistance range of candidate cacao plants.
Cacao↗
CaAlaAT1 catalyzes the alanine: 2-oxoglutarate aminotransferase reaction during the resistance response against Tobacco mosaic virus in hot pepper.
Hot pepper (Capsicum annuum L. cv. Bugang) plants exhibit a hypersensitive response (HR) upon infection by Tobacco mosaic virus (TMV) pathotype P0. To elucidate molecular mechanism that underlies this resistance, hot pepper cv. Bugang leaves were inoculated with TMV-P0 and genes specifically up-regulated during the HR were isolated by differential screening. One of the clones, CaAlaAT1 encoding a putative alanine aminotransferase (EC 2.6.1.2) exhibited organ-specific expression pattern and the transcript accumulated abundantly in red (ripe) fruit tissues. CaAlaAT1 transcript was also induced in older leaves during senescence. The expression of CaAlaAT1 gene was increased in the incompatible interaction with TMV-P0 but was not in the compatible interaction with TMV-P1.2. When a strain of Xanthomonas campestris pv. vesicatoria (Xcv) carrying an AvrBs2 gene was infiltrated into the leaves of a pepper cv. ECW 20R carrying Bs2 resistance gene, a marked induction and maintenance of CaAlaAT1 gene expression was observed. The expression of CaAlaAT1 gene was triggered by salicylic acid (SA) and ethylene but not by methyl jasmonate (MeJA). CaAlaAT1 seemed to be localized mostly at the cytosol from the polyethylene glycol (PEG)-mediated transformation experiment. CaAlaAT1 seemed to catalyze alanine: 2-oxoglutarate aminotransferase (AKT) reaction, which was a main activity among the four activities in vitro, during the resistance response against TMV in hot pepper. These results suggest that CaAlaAT1, a protein known to be involved in metabolic reactions, might be one of the components in the plant's defense signal pathway against pathogens.
Alanine Transaminase↗
A large scale analysis of resistance gene homologues in Arachis.
Arachis hypogaea L., commonly known as the peanut or groundnut, is an important and widespread food legume. Because the crop has a narrow genetic base, genetic diversity in A. hypogaea is low and it lacks sources of resistance to many pests and diseases. In contrast, wild diploid Arachis species are genetically diverse and are rich sources of disease resistance genes. The majority of known plant disease resistance genes encode proteins with a nucleotide binding site domain (NBS). In this study, degenerate PCR primers designed to bind to DNA regions encoding conserved motifs within this domain were used to amplify NBS-encoding regions from Arachis spp. The Arachis spp. used were A. hypogaea var. Tatu and wild species that are known to be sources of disease resistance: A. cardenasii, A. duranensis, A. stenosperma and A. simpsonii. A total of 78 complete NBS-encoding regions were isolated, of which 63 had uninterrupted ORFs. Phylogenetic analysis of the Arachis NBS sequences derived in this study and other NBS sequences from Arabidopsis thaliana, Medicago trunculata, Glycine max, Lotus japonicus and Phaseolus vulgaris that are available in public databases This analysis indicates that most Arachis NBS sequences fall within legume-specific clades, some of which appear to have undergone extensive copy number expansions in the legumes. In addition, NBS motifs from A. thaliana and legumes were characterized. Differences in the TIR and non-TIR motifs were identified. The likely effect of these differences on the amplification of NBS-encoding sequences by PCR is discussed.
Arabidopsis↗
Organization and evolution of resistance gene analogs in peanut.
The scarcity of genetic polymorphism in Arachis hypogaea (peanut), as in other monophyletic polyploid species, makes it especially vulnerable to nematode, bacterial, fungal, and viral pathogens. Although no disease resistance genes have been cloned from peanut itself, the conserved motifs in cloned resistance genes from other plant species provide a means to isolate and analyze similar genes from peanut. To survey the number, diversity, evolutionary history, and genomic organization of resistance gene-like sequences in peanut, we isolated 234 resistance gene analogs (RGAs) by using primers designed from conserved regions of different classes of resistance genes including NBS-LRR, and LRR-TM classes. Phylogenetic and sequence analyses were performed to explore evolutionary relationships both among peanut RGAs and with orthologous genes from other plant taxa. Fifty-six overgos designed from the RGA sequences on the basis of their phyletic association were applied to a peanut BAC library; 736 hybridizing BAC clones were fingerprinted and contigs were formed in order to gain insights into the genomic organization of these genes. All the fingerprinting gels were blotted and screened with the respective overgos in order to verify the authenticity of the hits from initial screens, and to explore the physical organization of these genes in terms of both copy number and distribution in the genome. As a result, we identified 250 putative resistance gene loci. A correlation was found between the phyletic positions of the sequences and their physical locations. The BACs isolated here will serve as a valuable resource for future applications, such as map-based cloning, and will help improve our understanding of the evolution and organization of these genes in the peanut genome.
Amino Acid Sequence↗
Genome-wide investigation on the genetic variations of rice disease resistance genes.
Exploitation of plant disease resistance (R) gene in breeding programs has been proven to be the most efficient strategy for coping with the threat of pathogens. An understanding of R-gene variation is the basis for this strategy. Here we report a genome-wide investigation on the variation of NBS-LRR-encoding genes, the common type of R genes, between two sequenced rice genomes, Oryza sativa L. var. Nipponbare and 93-11. We show that the allelic nucleotide diversity in 65.0% of 397 least-divergent pairs is not high (0.344% on average), while the remaining 35% display a greater diversity (5.4% on average). The majority of conserved R genes is single-copy and/or located as a singleton. The clustered, particularly the complex-clustered, R-genes contribute greatly to the rich genetic variation. Surprisingly only 11.2% of R-genes have remarkably high ratios of non-synonymous to synonymous rates, which is much less than the 17.4% observed between Arabidopsis genomes. Noticeable "artificially selective sweeping" could be detected in a large proportion of the conserved R-genes, a scenario described in the "arms race" co-evolutionary model. Based on our study, a variation pattern of R-genes is proposed and confirmed by the analysis of R-genes from other rice lines, indicating that the observed variation pattern may be common in all rice lines.
Genes, Plant↗
Induction of broadly neutralizing antibodies against measles virus mutants using a polyepitope vaccine strategy.
Chimeric molecules expressing multiple copies of the loop-forming hemagglutinin noose epitope (designated as "L"; aa386-400), a protective B cell epitope of the measles virus were generated by recombinant technology. The recombinant polyepitope [L4T4]2 combining two sets of four repeats of the L epitope and with two sets of four repeats of the human promiscuous T cell epitope of tetanus toxoid ("T", tt830-844) was produced in transgenic carrot plants. After intraperitoneal immunization of mice with plant membrane extract, sera neutralized all wild-type viruses. In a modified plaque reduction neutralization assay based on CD150-transfected Vero cells anti-[L4T4]2 sera neutralized all field isolates, irrespective of mutations in the L epitope. Even viruses with a mutation in the contact residues of a neutralizing L-specific monoclonal antibody or two mutations in other positions of the epitope were equally sensitive to neutralization. These results suggest that the multiple copies of the L epitope fold into different conformations that induce a repertoire of B cells diverse enough to overcome the genetic diversity of field viruses.
Amino Acid Sequence↗