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Exploration of cotton leaf curl virus resistance genes and their screening in Gossypium arboreum by targeting resistance gene analogues.

ABSTRACT: Cotton leaf curl virus (CLCuV) disease is one of the major limiting factors in cotton production, particularly in widely cultivated Gossypium hirsutum varieties that are susceptible to attack by this virus. Several approaches have been employed to explore putative resistance genes in another cotton species, G. arboreum. However, the exact mechanisms conferring disease resistance in cotton are still unknown. In the current study, we used various approaches to identify possible resistance genes against CLCuV infection. We report the identification and isolation of a set of genes involved in the resistance response to viral infestation. PCR products containing genomic DNA gave multiple amplifications with a single primer in most reactions, and 38 fragments were cloned from G. arboreum and G. hirsutum. The sequences of cloned fragments belonged to various pathway genes and uncharacterized proteins. However, five amplified fragments (RM1, RM6, RM8, RM12 and RM31) showed similarity with R genes. Maximum homology (94 %) was observed with G. raimondii toll/interleukin receptor-like protein. BLAST search showed the homology of all resistance gene analogues (RGAs) with more than one chromosome, and multiple hits were observed on each chromosome for each RGA. Expression analysis through RT-PCR identified variable expression levels of the different RGAs in all tested genotypes. The expression level of RGAs differed between symptomatic and asymptomatic plants, with the exception of RGA 395, whose expression level was the same in both diseased and healthy plants. Knowledge of the interaction of these genes with various cotton pathogens could be utilized to improve the resistance of susceptible G. hirsutum and other plant species.

SUBMITTER: Mushtaq R

PROVIDER: S-EPMC6247833 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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Exploration of cotton leaf curl virus resistance genes and their screening in Gossypium arboreum by targeting resistance gene analogues.

Mushtaq Rakhshanda R Shahzad Khurram K Mansoor Shahid S Shah Zahid Hussain ZH Alsamadany Hameed H Mujtaba Tahir T Al-Zahrani Yahya Y Alzahrani Hind A S HAS Ahmed Zaheer Z Bashir Aftab A

AoB PLANTS 20181016 6

Cotton leaf curl virus (CLCuV) disease is one of the major limiting factors in cotton production, particularly in widely cultivated Gossypium hirsutum varieties that are susceptible to attack by this virus. Several approaches have been employed to explore putative resistance genes in another cotton species, G. arboreum. However, the exact mechanisms conferring disease resistance in cotton are still unknown. In the current study, we used various approaches to identify possible resis ...[more]

PMID: 30487965

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Project description:BackgroundWhitefly-transmitted geminiviruses (family Geminiviridae, genus Begomovirus) are major limiting factors for the production of numerous dicotyledonous crops throughout the warmer regions of the world. In the Old World a small number of begomoviruses have genomes consisting of two components whereas the majority have single-component genomes. Most of the monopartite begomoviruses associate with satellite DNA molecules, the most important of which are the betasatellites. Cotton leaf curl disease (CLCuD) is one of the major problems for cotton production on the Indian sub-continent. Across Pakistan, CLCuD is currently associated with a single begomovirus (Cotton leaf curl Burewala virus [CLCuBuV]) and the cotton-specific betasatellite Cotton leaf curl Multan betasatellite (CLCuMuB), both of which have recombinant origins. Surprisingly, CLCuBuV lacks C2, one of the genes present in all previously characterized begomoviruses. Virus-specific transcripts have only been mapped for few begomoviruses, including one monopartite begomovirus that does not associate with betasatellites. Similarly, the transcripts of only two betasatellites have been mapped so far. The study described has investigated whether the recombination/mutation events involved in the evolution of CLCuBuV and its associated CLCuMuB have affected their transcription strategies.ResultsThe major transcripts of CLCuBuV and its associated betasatellite (CLCuMuB) from infected Nicotiana benthamiana plants have been determined. Two complementary-sense transcripts of ~1.7 and ~0.7 kb were identified for CLCuBuV. The ~1.7 kb transcript appears similar in position and size to that of several begomoviruses and likely directs the translation of C1 and C4 proteins. Both complementary-sense transcripts can potentially direct the translation of C2 and C3 proteins. A single virion-sense transcript of ~1 kb, suitable for translation of the V1 and V2 genes was identified. A predominant complementary-sense transcript was also confirmed for the betasatellite.ConclusionsOverall, the transcription of CLCuBuV and the recombinant CLCuMuB is equivalent to earlier mapped begomoviruses/betasatellites. The recombination events that featured in the origins of these components had no detectable effects on transcription. The transcripts spanning the mutated C2 gene showed no evidence for involvement of splicing in restoring the ability to express intact C2 protein.

Project description:Cotton is a commercial and economically important crop that generates billions of dollars in annual revenue worldwide. However, cotton yield is affected by a sap-sucking insect Bemisia tabaci (whitefly), and whitefly-borne cotton leaf curl disease (CLCuD). The causative agent of devastating CLCuD is led by the viruses belonging to the genus Begomovirus (family Geminiviridae), collectively called cotton leaf curl viruses. Unfortunately, the extensively cultivated cotton (Gossypium hirsutum) species are highly susceptible and vulnerable to CLCuD. Yet, the concomitant influence of whitefly and CLCuD on the susceptible G. hirsutum transcriptome has not been interpreted. In the present study we have employed an RNA Sequencing (RNA-Seq) transcriptomics approach to explore the differential gene expression in susceptible G. hirsutum variety upon infection with viruliferous whiteflies. Comparative RNA-Seq of control and CLCuD infected plants was done using Illumina HiSeq 2500. This study yielded 468 differentially expressed genes (DEGs). Among them, we identified 220 up and 248 downregulated DEGs involved in disease responses and pathogen defense. We selected ten genes for downstream RT-qPCR analyses on two cultivars, Karishma and MNH 786 that are susceptible to CLCuD. We observed a similar expression pattern of these genes in both susceptible cultivars that was also consistent with our transcriptome data further implying a wider application of our global transcription study on host susceptibility to CLCuD. We next performed weighted gene co-expression network analysis that revealed six modules. This analysis also identified highly co-expressed genes as well as 55 hub genes that co-express with ≥ 50 genes. Intriguingly, most of these hub genes are shown to be downregulated and enriched in cellular processes. Under-expression of such highly co-expressed genes suggests their roles in favoring the virus and enhancing plant susceptibility to CLCuD. We also discuss the potential mechanisms governing the establishment of disease susceptibility. Overall, our study provides a comprehensive differential gene expression analysis of G. hirsutum under whitefly-mediated CLCuD infection. This vital study will advance the understanding of simultaneous effect of whitefly and virus on their host and aid in identifying important G. hirsutum genes which intricate in its susceptibility to CLCuD.

Dataset Information

Exploration of cotton leaf curl virus resistance genes and their screening in Gossypium arboreum by targeting resistance gene analogues.

Publications

Exploration of cotton leaf curl virus resistance genes and their screening in <i>Gossypium arboreum</i> by targeting resistance gene analogues.

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