Genome-Wide Identification and Analysis of CC-NBS-LRR Family in Response to Downy Mildew and Black Rot in Chinese Cabbage.
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ABSTRACT:
SUBMITTER: Liu Y
PROVIDER: S-EPMC8074028 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Powdery mildew caused by Blumeria graminis f. sp. tritici (Bgt) is a destructive disease of wheat throughout the world. Host resistance is considered the most sustainable way to control this disease. Powdery mildew resistance gene Pm2b was mapped to the same genetic interval with Pm2a and PmCH1357 cloned previously, but showed different resistance spectra from them, indicating that they might be caused by different resistance genes or alleles. In this study, Pm2b was delimited to a 1.64 Mb physical interval using a large segregating population containing 4,354 F2:3 families of resistant parent KM2939 and susceptible cultivar Shimai 15. In this interval, TraesCS5D03G0111700 encoding the coiled-coil nucleotide-binding site leucine-rich repeat protein (CC-NBS-LRR) was determined as the candidate gene of Pm2b. Silencing by barley stripe mosaic virus-induced gene silencing (BSMV-VIGS) technology and two independent mutants analysis in KM2939 confirmed the candidate gene TraesCS5D03G0111700 was Pm2b. The sequence of Pm2b was consistent with Pm2a/PmCH1357. Subcellular localization showed Pm2b was located on the cell nucleus and plasma membrane. Pm2b had the highest expression level in leaves and was rapidly up-regulated after inoculating with Bgt isolate E09. The yeast two-hybrid (Y2H) and luciferase complementation imaging assays (LCI) showed that PM2b could self-associate through the NB domain. Notably, we identified PM2b interacting with the transcription factor TaWRKY76-D, which depended on the NB domain of PM2b and WRKY domain of TaWRKY76-D. TaWRKY76-D negatively regulated the resistance to powdery mildew in wheat. The specific KASP marker K529 could take the advantage of high-throughput and high-efficiency for detecting Pm2b and be useful in molecular marker assisted-selection breeding. In conclusion, cloning and disease resistance mechanism analysis of Pm2b provided an example to emphasize a need of the molecular isolation of resistance genes, which has implications in marker assisted wheat breeding.
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Project description:Rye (Secale cereale), a valuable relative of wheat, contains abundant powdery mildew resistance (Pm) genes. Using physical mapping, transcriptome sequencing, barley stripe mosaic virus-induced gene silencing, ethyl methane sulfonate mutagenesis, and stable transformation, we isolated and validated two coiled-coil, nucleotide-binding site and leucine-rich repeat (CC-NBS-LRR) alleles, PmTR1 and PmTR3, located on rye chromosome 6RS from different triticale lines. PmTR1 confers age-related resistance starting from the three-leaf stage, whereas its allele, PmTR3, confers typical all-stage resistance, which may be associated with their differential gene expression patterns. Overexpression in Nicotiana benthamiana showed that the CC, CC-NBS, and CC-LRR fragments of PMTR1 induce cell death, whereas in PMTR3 the CC and full-length fragments perform this function. Luciferase complementation imaging and pull-down assays revealed distinct interaction activities between the CC and NBS fragments. Our study elucidates two novel rye-derived Pm genes and their derivative germplasm resources and provides novel insights into the mechanism of age-related resistance, which can aid the improvement of resistance against wheat powdery mildew.
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Project description:Brassica downy mildew, a severe disease caused by Hyaloperonospora brassicae, can cause enormous economic losses in Chinese cabbage (Brassica rapa L. ssp. pekinensis) production. Although some research has been reported recently concerning the underlying resistance to this disease, no studies have identified or characterized long noncoding RNAs involved in this defense response. In this study, using high-throughput RNA sequencing, we analyzed the disease-responding mRNAs and long noncoding RNAs in two resistant lines (T12-19 and 12-85) and one susceptible line (91-112). Clustering and Gene Ontology analysis of differentially expressed genes (DEGs) showed that more DEGs were involved in the defense response in the two resistant lines than in the susceptible line. Different expression patterns and proposed functions of differentially expressed long noncoding RNAs among T12-19, 12-85, and 91-112 indicated that each has a distinct disease response mechanism. There were significantly more cis- and trans-functional long noncoding RNAs in the resistant lines than in the susceptible line, and the genes regulated by these RNAs mostly participated in the disease defense response. Furthermore, we identified a candidate resistance-related long noncoding RNA, MSTRG.19915, which is a long noncoding natural antisense transcript of a MAPK gene, BrMAPK15. Via an agroinfiltration-mediated transient overexpression system and virus-induced gene silencing technology, BrMAPK15 was indicated to have a greater ability to defend against pathogens. MSTRG.19915-silenced seedlings showed enhanced resistance to downy mildew, probably because of the upregulated expression of BrMAPK15. This research identified and characterized long noncoding RNAs involved in resistance to downy mildew, laying a foundation for future in-depth studies of disease resistance mechanisms in Chinese cabbage.
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Project description:Hybrid lethality forms a reproductive barrier that has been found in many eukaryotes. Most cases follow the Bateson-Dobzhansky-Muller genetic incompatibility model and involve two or more loci. In this study, we demonstrate that a coiled-coil nucleotide-binding site leucine-rich repeat (CC-NBS-LRR) gene is the causal gene underlying the Le4 locus for interspecific hybrid lethality between Gossypium barbadense and G. hirsutum (cotton). Silencing this CC-NBS-LRR gene can restore F1 plants from a lethal to a normal phenotype. A total of 11 099 genes were differentially expressed between the leaves of normal and lethal F1 plants, of which genes related to autoimmune responses were highly enriched. Genes related to ATP-binding and ATPase were up-regulated before the lethal syndrome appeared; this may result in the conversion of Le4 into an active state and hence trigger immune signals in the absence of biotic/abiotic stress. We discuss our results in relation to the evolution and domestication of Sea Island cottons and the molecular mechanisms of hybrid lethality associated with autoimmune responses. Our findings provide new insights into reproductive isolation and may benefit cotton breeding.
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Project description:Grapevine downy mildew (DM) is a destructive oomycete disease of viticulture worldwide. MrRPV1 is a typical TIR-NBS-LRR type DM disease resistance gene cloned from the wild North American grapevine species Muscadinia rotundifolia. However, the molecular basis of resistance mediated by MrRPV1 remains poorly understood. Downy mildew-susceptible Vitis vinifera cv. Shiraz was transformed with a genomic fragment containing MrRPV1 to produce DM-resistant transgenic Shiraz lines. Comparative transcriptome analysis was used to compare the transcriptome profiles of the resistant and susceptible genotypes after DM infection. Transcriptome modulation during the response to P. viticola infection was more rapid, and more genes were induced in MrRPV1-transgenic Shiraz than in wild-type plants. In DM-infected MrRPV1-transgenic plants, activation of genes associated with Ca2+ release and ROS production was the earliest transcriptional response. Functional analysis of differentially expressed genes revealed that key genes related to multiple phytohormone signaling pathways and secondary metabolism were highly induced during infection. Coexpression network and motif enrichment analysis showed that WRKY and MYB transcription factors strongly coexpress with stilbene synthase (VvSTS) genes during defense against P. viticola in MrRPV1-transgenic plants. Taken together, these findings indicate that multiple pathways play important roles in MrRPV1-mediated resistance to downy mildew.
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Project description:The NBS-LRR (NLR) gene family plays a pivotal role in regulating disease defense response in plants. Cucumber is one of the most important vegetable crops in the world, and various plant diseases, including powdery mildew (PM), cause severe losses in both cucumber productivity and quality annually. To characterize and understand the role of the CC-NBS-LRR(CNL) family of genes in disease defense response in cucumber plants, we performed bioinformatical analysis to characterize these genes systematically. We identified 33 members of the CNL gene family in cucumber plants, and they are distributed on each chromosome with chromosome 4 harboring the largest cluster of five different genes. The corresponding CNL family member varies in the number of amino acids and exons, molecular weight, theoretical isoelectric point (pI) and subcellular localization. Cis-acting element analysis of the CNL genes reveals the presence of multiple phytohormone, abiotic and biotic responsive elements in their promoters, suggesting that these genes might be responsive to plant hormones and stress. Phylogenetic and synteny analysis indicated that the CNL proteins are conserved evolutionarily in different plant species, and they can be divided into four subfamilies based on their conserved domains. MEME analysis and multiple sequence alignment showed that conserved motifs exist in the sequence of CNLs. Further DNA sequence analysis suggests that CsCNL genes might be subject to the regulation of different miRNAs upon PM infection. By mining available RNA-seq data followed by real-time quantitative PCR (qRT-PCR) analysis, we characterized expression patterns of the CNL genes, and found that those genes exhibit a temporospatial expression pattern, and their expression is also responsive to PM infection, ethylene, salicylic acid, and methyl jasmonate treatment in cucumber plants. Finally, the CNL genes targeted by miRNAs were predicted in cucumber plants. Our results in this study provided some basic information for further study of the functions of the CNL gene family in cucumber plants.
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Project description:Verticillium wilt (VW) is a destructive disease in cotton caused by Verticillium dahliae and has a significant impact on yield and quality. In the absence of safe and effective chemical control, VW is difficult to manage. Thus, at present, developing resistant varieties is the most economical and effective method of controlling Verticillium wilt of cotton. The CC-NBS-LRR (CNL) gene family is an important class of plant genes involved in disease resistance. This study identified 141 GbCNLs in Gossypium barbadense genome, with 37.5% (53 genes) GbCNLs enriched in 12 gene clusters (GC01-GC12) based on gene distribution in the chromosomes. Especially, seven GbCNLs from two largest clusters (GC11 and GC12) were significantly upregulated in the resistant cultivar (Hai No. 7124) and the susceptible (Giza No. 57). Virus-induced gene silencing of GbCNL130 in G. barbadense, one typical gene in the gene cluster 12 (GC12), significantly altered the response to VW, compromising plant resistance to V. dahliae. In contrast, GbCNL130 overexpression significantly increased the resistance to VW in the wild-type Arabidopsis thaliana. Based on our research findings presented here, we conclude that GbCNL130 promotes resistance to VW by activating the salicylic acid (SA)-dependent defense response pathway resulting in strong accumulation of reactive oxygen species and upregulation of pathogenesis-related (PR) genes. In conclusion, our study resulted in the discovery of a new CNL resistance gene in cotton, GbCNL130, that confers resistance to VW across different hosts.
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Project description:Hexaploid wheat displays limited genetic variation. As a direct A and B genome donor of hexaploid wheat, tetraploid wheat represents an important gene pool for cultivated bread wheat. Many disease resistant genes express conserved domains of the nucleotide-binding site and leucine-rich repeats (NBS-LRR). In this study, we isolated a CC-NBS-LRR gene locating on chromosome 7B from durum wheat variety Italy 363, and designated it TdRGA-7Ba. Its open reading frame was 4014 bp, encoding a 1337 amino acid protein with a complete NBS domain and 18 LRR repeats, sharing 44.7% identity with the PM3B protein. TdRGA-7Ba expression was continuously seen at low levels and was highest in leaves. TdRGA-7Ba has another allele TdRGA-7Bb with a 4 bp deletion at position +1892 in other cultivars of tetraploid wheat. In Ae. speltoides, as a B genome progenitor, both TdRGA-7Ba and TdRGA-7Bb were detected. In all six species of hexaploid wheats (AABBDD), only TdRGA-7Bb existed. Phylogenic analysis showed that all TdRGA-7Bb type genes were grouped in one sub-branch. We speculate that TdRGA-7Bb was derived from a TdRGA-7Ba mutation, and it happened in Ae. speltoides. Both types of TdRGA-7B participated in tetraploid wheat formation. However, only the TdRGA-7Bb was retained in hexaploid wheat.
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Project description:Key messagePh-3 is the first cloned tomato gene for resistance to late blight and encodes a CC-NBS-LRR protein. Late blight, caused by Phytophthora infestans, is one of the most destructive diseases in tomato. The resistance (R) gene Ph-3, derived from Solanum pimpinellifolium L3708, provides resistance to multiple P. infestans isolates and has been widely used in tomato breeding programmes. In our previous study, Ph-3 was mapped into a region harbouring R gene analogues (RGA) at the distal part of long arm of chromosome 9. To further narrow down the Ph-3 interval, more recombinants were identified using the flanking markers G2-4 and M8-2, which defined the Ph-3 gene to a 26 kb region according to the Heinz1706 reference genome. To clone the Ph-3 gene, a bacterial artificial chromosome (BAC) library was constructed using L3708 and one BAC clone B25E21 containing the Ph-3 region was identified. The sequence of the BAC clone B25E21 showed that only one RGA was present in the target region. A subsequent complementation analysis demonstrated that this RGA, encoding a CC-NBS-LRR protein, was able to complement the susceptible phenotype in cultivar Moneymaker. Thus this RGA was considered the Ph-3 gene. The predicted Ph-3 protein shares high amino acid identity with the chromosome-9-derived potato resistance proteins against P. infestans (Rpi proteins).
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Project description:Eucalyptus grandis is a commercially important hardwood species and is known to be susceptible to a number of pests and pathogens. Determining mechanisms of defense is therefore a research priority. The published genome for E. grandis has aided the identification of one important class of resistance (R) genes that incorporate nucleotide binding sites and leucine-rich repeat domains (NBS-LRR). Using an iterative search process we identified NBS-LRR gene models within the E. grandis genome. We characterized the gene models and identified their genomic arrangement. The gene expression patterns were examined in E. grandis clones, challenged with a fungal pathogen (Chrysoporthe austroafricana) and insect pest (Leptocybe invasa). One thousand two hundred and fifteen putative NBS-LRR coding sequences were located which aligned into two large classes, Toll or interleukin-1 receptor (TIR) and coiled-coil (CC) based on NB-ARC domains. NBS-LRR gene-rich regions were identified with 76% organized in clusters of three or more genes. A further 272 putative incomplete resistance genes were also identified. We determined that E. grandis has a higher ratio of TIR to CC classed genes compared to other woody plant species as well as a smaller percentage of single NBS-LRR genes. Transcriptome profiles indicated expression hotspots, within physical clusters, including expression of many incomplete genes. The clustering of putative NBS-LRR genes correlates with differential expression responses in resistant and susceptible plants indicating functional relevance for the physical arrangement of this gene family. This analysis of the repertoire and expression of E. grandis putative NBS-LRR genes provides an important resource for the identification of novel and functional R-genes; a key objective for strategies to enhance resilience.
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