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Summary Improving genetic resistance is a preferred method to manage Verticillium wilt of cotton and other hosts. Identifying host resistance is difficult because of the dearth of resistance genes against this pathogen. Previously, a novel candidate gene involved in Verticillium wilt resistance was identified by a genome‐wide association study using a panel of Gossypium hirsutum accessions. In this study, we cloned the candidate resistance gene from cotton that encodes a protein sharing homology with the TIR‐NBS‐LRR receptor‐like defence protein DSC1 in Arabidopsis thaliana (hereafter named GhDSC1). GhDSC1 expressed at higher levels in response to Verticillium wilt and jasmonic acid (JA) treatment in resistant cotton cultivars as compared to susceptible cultivars and its product was localized to nucleus. The transfer of GhDSC1 to Arabidopsis conferred Verticillium resistance in an A. thaliana dsc1 mutant. This resistance response was associated with reactive oxygen species (ROS) accumulation and increased expression of JA‐signalling‐related genes. Furthermore, the expression of GhDSC1 in response to Verticillium wilt and JA signalling in A. thaliana displayed expression patterns similar to GhCAMTA3 in cotton under identical conditions, suggesting a coordinated DSC1 and CAMTA3 response in A. thaliana to Verticillium wilt. Analyses of GhDSC1 sequence polymorphism revealed a single nucleotide polymorphism (SNP) difference between resistant and susceptible cotton accessions, within the P‐loop motif encoded by GhDSC1. This SNP difference causes ineffective activation of defence response in susceptible cultivars. These results demonstrated that GhDSC1 confers Verticillium resistance in the model plant system of A. thaliana, and therefore represents a suitable candidate for the genetic engineering of Verticillium wilt resistance in cotton.

Plant disease resistance genes (R genes) encode proteins that function to monitor signals indicating pathogenic infection, thus playing a critical role in the plant's defense system. Although many studies have been performed to explore the functional details of these important genes, their origin and evolutionary history remain unclear. In this study, focusing on the largest group of R genes, the nucleotide-binding site–leucinerich repeat (NBS-LRR) genes, we conducted an extensive genome-wide survey of 38 representative model organisms and obtained insights into the evolutionary stage and timing of NBS-LRR genes. Our data show that the two major domains, NBS and LRR, existed before the split of prokaryotes and eukaryotes but their fusion was observed only in land plant lineages. The Toll/interleukin-1 receptor (TIR) class of NBS-LRR genes probably had an earlier origin than its nonTIR counterpart. The similarities of the innate immune systems of plants and animals are likely to have been shaped by convergent evolution after their independent origins. Our findings start to unravel the evolutionary history of these important genes from the perspective of comparative genomics and also highlight the important role of reorganizing pre-existing building blocks in generating evolutionary novelties.

Citrus is a highly important global fruit crop that faces significant challenges from citrus Huanglongbing (HLB). The nucleotide binding site leucine-rich repeat ( NBS-LRR ) gene family plays a central role in plant immunity. However, comprehensive studies on the function of NBS-LRR genes in HLB resistance or tolerance remain limited. In this study, we identified 278 NBS-LRR genes from Citrus clementina . These genes exhibit 7 domain architectures. Physicochemical analyses revealed significant subfamily divergence in length (91-2101 aa), molecular weight (10.57-237.18 kDa), and isoelectric point (pI, 4.61–9.35). Over 82% of proteins localize to the nucleus/cytoplasm, indicating roles in signal transduction and transcriptional regulation. NBS-LRR genes cluster non-randomly among chromosomes, forming tandem repeats on chromosomes 3, 5, and 7. Phylogenetic analysis grouped the NBS-LRR family into 5 clades. Conserved domain analysis identified 10 motifs; Motif 1 and 2 are highly conserved, potentially involved in pathogen recognition. Promoter analysis detected 36 cis-acting elements linked to diverse processes (e.g., hormone/stress response). Synteny analysis revealed 9 genes are orthologous genes of Arabidopsis thaliana . Crucially, transcriptome data showed that significantly higher NBS-LRR expression in Huanglongbing-infected plants, pinpointing candidate gene Ciclev10018143m.1 . This study provides a foundation for citrus disease resistance breeding and candidate genes for functional validation.

Ascochyta blight is one of the major diseases of chickpea worldwide. The genetic resistance to ascochyta blight in chickpea is complex and governed by multiple QTLs. However, the molecular mechanism of quantitative disease resistance to ascochyta blight and the genes underlying these QTLs are still unknown. Most often disease resistance is determined by resistance (R) genes. The most predominant R-genes contain nucleotide binding site and leucine rich repeat (NBS-LRR) domains. A total of 121 NBS-LRR genes were identified in the chickpea genome. Ninety-eight of these genes contained all essential conserved domains while 23 genes were truncated. The NBS-LRR genes were grouped into eight distinct classes based on their domain architecture. Phylogenetic analysis grouped these genes into two major clusters based on their structural variation, the first cluster with toll or interleukin-1 like receptor (TIR) domain and the second cluster either with or without a coiled-coil domain. The NBS-LRR genes are distributed unevenly across the eight chickpea chromosomes and nearly 50% of the genes are present in clusters. Thirty of the NBS-LRR genes were co-localized with nine of the previously reported ascochyta blight QTLs and were tested as potential candidate genes for ascochyta blight resistance. Expression pattern of these genes was studied in two resistant (CDC Corinne and CDC Luna) and one susceptible (ICCV 96029) genotypes at different time points after ascochyta blight infection using real-time quantitative PCR. Twenty-seven NBS-LRR genes showed differential expression in response to ascochyta blight infection in at least one genotype at one time point. Among these 27 genes, the majority of the NBS-LRR genes showed differential expression after inoculation in both resistant and susceptible genotypes which indicates the involvement of these genes in response to ascochyta blight infection. Five NBS-LRR genes showed genotype specific expression. Our study provides a new insight of NBS-LRR gene family in chickpea and the potential involvement of NBS-LRR genes in response to ascochyta blight infection.

Background: The NBS-LRR gene family plays a critical role in plant disease resistance and is considered a key determinant of plant immune responses. Research on the NBS-LRR gene family has grown rapidly, with significant progress driven by advances of molecular biology techniques. However, to date, there has been no systematic identification of NBS-LRR genes in Nicotiana species. Methods: In this study, we systematically characterized the NBS gene families in three Nicotiana genomes, investigated the evolution and environmental selection during the species formation, and explored the key NBS genes involved in disease resistance. Results: Results showed that 1226 NBS genes are present across the three Nicotiana genomes, and 76.62% of the members in Nicotiana tabacum could be traced back to their parental genomes. In addition, whole-genome duplication was found to contribute significantly to the expansion of NBS gene families. In addition, many NBS genes associated with disease resistance were identified, including one multi-disease resistance gene. Conclusions: This study provides new insights into the formation of NBS gene families in Nicotiana and offers new clues for understanding plant immunity.

During plant evolution, nucleotide-binding sites (NBS) and leucine-rich repeat (LRR) genes have made significant contributions to plant disease resistance. With many high-quality plant genomes sequenced, identification and comprehensive analyses of NBS-LRR genes at whole genome level are of great importance to understand and utilize them. In this study, we identified the NBS-LRR genes of 23 representative species at whole genome level, and researches on NBS-LRR genes of four monocotyledonous grass species, Saccharum spontaneum, Saccharum officinarum, Sorghum bicolor and Miscanthus sinensis, were focused. We found that whole genome duplication, gene expansion, and allele loss could be factors affecting the number of NBS-LRR genes in the species, and whole genome duplication is likely to be the main cause of the number of NBS-LRR genes in sugarcane. Meanwhile, we also found a progressive trend of positive selection on NBS-LRR genes. These studies further elucidated the evolutionary pattern of NBS-LRR genes in plants. Transcriptome data from multiple sugarcane diseases revealed that more differentially expressed NBS-LRR genes were derived from S. spontaneum than from S. officinarum in modern sugarcane cultivars, and the proportion was significantly higher than the expected. This finding reveals that S. spontaneum has a greater contribution to disease resistance for modern sugarcane cultivars. In addition, we observed allelespecific expression of seven NBS-LRR genes under leaf scald, and 125 NBS-LRR genes responding to multiple diseases were identified. Finally, we built a plant NBS-LRR gene database to facilitate subsequent analysis and use of NBSLRR genes obtained here. In conclusion, this study complemented and completed the research of plant NBS-LRR genes, and discussed how NBS-LRR genes responding to sugarcane diseases, which provided a guide and genetic resources for further research and utilization of NBS-LRR genes.

Alternaria brown spot, caused by the tangerine pathotype of Alternaria alternata, is a destructive fungal disease affecting citrus production worldwide. Nucleotide-binding site-leucine-rich repeat (NBS-LRR) genes constitute a major class of plant immune receptors; however, their genome-wide characteristics and potential association with Alternaria brown spot resistance loci in citrus remain poorly understood. In this study, we performed a comprehensive genome-wide identification and comparative analysis of NBS-LRR genes across representative citrus species. A total of 417 and 326 NBS-LRR genes were identified in Citrus reticulata and Citrus clementina, respectively, and were classified into NL, CNL, TNL, and RNL subfamilies based on domain architecture. Phylogenetic reconstruction, gene structure analysis, conserved motif composition, chromosomal distribution, synteny relationships, and promoter cis-element profiling collectively revealed considerable structural variation and lineage-specific expansion of the NBS-LRR gene family in citrus genomes. By integrating previously reported quantitative trait locus (QTL) data for Alternaria brown spot, we identified several NBS-LRR genes located within a resistance-associated genomic interval on chromosome 3. Among these, a candidate gene, designated LRR2, exhibited differential transcriptional responses upon pathogen inoculation and displayed distinct sequence variations between citrus genotypes. Structural modeling and molecular docking analyses suggested potential binding interfaces between LRR2 and multiple host-selective toxins, although the biological relevance of these interactions requires further experimental validation. Subcellular localization assays in Nicotiana benthamiana showed that LRR2 is distributed in both the nucleus and cytoplasm. Notably, transient overexpression of LRR2 triggered hypersensitive response-like cell death and H2O2 accumulation. Collectively, this study provides a comprehensive overview of the citrus NBS-LRR gene family and presents a multifaceted characterization of a QTL-anchored candidate gene. These findings establish a genomic and molecular framework for further functional investigations of citrus–Alternaria interactions.

Most of the currently available disease resistance (R) genes have NBS (nucleotide-binding site) and LRR (leucine-rich-repeat) domain which belongs to the NBS-LRR gene family. The whole genome sequencing of Broussonetia papyrifera provides an important bioinformatics database for the study of the NBS-LRR gene family. In this study, 328 NBS-LRR family genes were identified and classified in B. papyrifera according to different classification schemes, where there are 92 N types, 47 CN type, 54 CNL type, 29 NL types, 55 TN type, and 51 TNL type. Subsequently, we conducted bioinformatics analysis of the NBS-LRR gene family. Classification, motif analysis of protein sequences, and phylogenetic tree studies of the NBS-LRR genes in B. papyrifera provide important basis for the functional study of NBS-LRR family genes. Additionally, we performed structural analysis of the chromosomal location, physicochemical properties, and sequences identified by genetic characterization. In addition, through the analysis of GO enrichment, it was found that NBS-LRR genes were involved in defense responses and were significantly enriched in biological stimulation, immune response, and abiotic stress. In addition, we found that Bp06g0955 was the most sensitive to low temperature and encoded the RPM1 protein by analyzing the low temperature transcriptome data of B. papyrifera. Quantitative results of gene expression after 48 h of Fusarium infection showed that Bp01g3293 increased 14 times after infection, which encodes RPM1 protein. The potential of NBS-LRR gene responsive to biotic and abiotic stresses can be exploited to improve the resistance of B. papyrifera.

NBS domain-containing sequences from the Agave tequilana transcriptome shotgun assembly were identified, characterized, and classified based on their physicochemical properties and motif structure, which resulted in a differential response to Lasiodiplodia sp. infection. Agave tequilana is an important crop in Mexico that is susceptible to many pathogens and adverse conditions. In plants, NBS-LRR genes are involved in the physiological response to pathogenic infection. Forty-six partial NBS-LRR sequences were identified in the transcriptome shotgun assembly and were classified into five subclasses (CNL, CN, NL, N, and L) belonging to the non-TIR class in the NBS-LRR family. The identified sequences encode functional NBS-LRR proteins based on physicochemical properties, gene structure and motif analysis, functional annotation, and gene ontology. Phylogenetic analysis showed that these genes were clustered into seven groups (Groups I-VII). These groups were under diversifying selection pressure (Ka/Ks rates < 1) except for Group V (Ka/Ks rate = 1.23) which formed more recently (9 Mya). Specific primers designed for Groups I, II and V showed that the expression response to pathogenic Lasiodiplodia strains varied among the different NBS-LRR gene groups. The highest NBS-LRR gene transcript induction was obtained at 48 h, and the expression peaks were preceded by an increase in the concentration of endogenous salicylic acid, which has been associated with the activation of some NBS-LRR genes, suggesting that each group may have a specific defence response function.

In this study, genome-wide identification, phylogenetic relationships, duplication time and selective pressure of the NBS-LRR genes, an important group of plant disease-resistance genes (R genes), were performed to uncover their genetic evolutionary patterns in the six Prunus species. A total of 1946 NBS-LRR genes were identified; specifically, 589, 361, 284, 281, 318, and 113 were identified in Prunus yedoensis, P. domestica, P. avium, P. dulcis, P. persica and P. yedoensis var. nudiflora, respectively. Two NBS-LRR gene subclasses, TIR-NBS-LRR (TNL) and non-TIR-NBS-LRR (non-TNL), were also discovered. In total, 435 TNL and 1511 non-TNL genes were identified and could be classified into 30/55/75 and 103/158/191 multi-gene families, respectively, according to three different criteria. Higher Ks and Ka/Ks values were detected in TNL gene families than in non-TNL gene families. These results indicated that the TNL genes had more members involved in relatively ancient duplications and were affected by stronger selection pressure than the non-TNL genes. In general, the NBS-LRR genes were shaped by species-specific duplications, and lineage-specific duplications occurred at recent and relatively ancient periods among the six Prunus species. Therefore, different duplicated copies of NBS-LRRs can resist specific pathogens and will provide an R-gene library for resistance breeding in Prunus species.