@article{XU2023, 
author = {FuChun XU and JingRuo ZHAO and ZhenNan ZHANG and GaiYuan HU and Lu LONG},
title = {Cloning and Functional Characterization of GhCPR5 in Disease Resistance of Gossypium hirsutum},
year = {2023},
journal = {Scientia Agricultura Sinica},
volume = {56},
number = {19},
pages = {3747-3758},
keywords = {cotton, disease resistance protein, Verticillium dahliae, Botrytis cinerea, gene expression},
url = {https://www.sciopen.com/article/10.3864/j.issn.0578-1752.2023.19.004},
doi = {10.3864/j.issn.0578-1752.2023.19.004},
abstract = {【Objective】This study on the function and mechanism of GhCPR5 in response to Verticillium dahliae (V. dahliae) and Botrytis cinerea (B. cinerea) in cotton analysed nucleotide sequence, protein structure, expression pattern, and biological function to provide a theoretical basis and genetic resources for cotton disease resistance and breeding mechanism research.【Method】GhCPR5 was identified from the unpublished transcriptome data of cotton responses to V. dahliae infection. The full-length coding sequence of GhCPR5 was amplified from upland cotton TM-1. The conserved domain, protein structure, and phylogenetic relationship of GhCPR5 and homologous genes were analysed using bioinformatics techniques. Real-time fluorescence quantitative PCR (qPCR) was used to analyse the GhCPR5 expression patterns in cotton roots, stems, leaves, ovules, fibres, and petals, and the induced expression of GhCPR5 by V. dahliae infection. The silencing fragments of GhCPR5 were amplified and inserted into the VIGS vector to generate the gene silencing construct TRV:CPR5. GhCPR5-silencing plants were created via Agrobacterium-mediated transformation methods. RT-PCR and qPCR were used to analyse the silencing efficiency of GhCPR5 in TRV:CPR5 plants. TRV:00 and TRV:CPR5 plants were inoculated with V. dahliae and B. cinerea, respectively, to analyse the difference in resistance in TRV:00 and TRV:CPR5 plants in response to pathogens. To analyse defence signal pathways involving GhCPR5, the expression levels of defence-related genes in TRV:00 and TRV:CPR5 plants were detected by qPCR.【Result】The GhCPR5 cloned from G. hirsutum TM-1 is 1 683 bp long and encodes a 560 amino acid protein. The relative molecular weight and isoelectric point of GhCPR5 are 62.883 kDa and 9.01, respectively. Multiple-sequence alignment and phylogenetic analysis showed that GhCPR5 is highly homologous with the CPR5 of Durio zibethinus and Theobroma cacao. Furthermore, the C-terminals of GhCPR5 and CPR5 of other species are highly conserved and contain 4-5 transmembrane domains. The GhCPR5 expression level was induced by V. dahliae infection and is highest in leaves and lowest in stems. Under normal conditions, no significant developmental differences were observed between the GhCPR5-silencing plants, TRV:CPR5, and the TRV:00 control plants. After inoculation with V. dahliae, the rate of disease and the disease index of TRV:CPR5 plants were significantly higher than those of the control plants. Analysis of detached leaves inoculated with V. dahliae and B. cinerea and lactophenol-trypan blue staining showed that the lesions on the leaves of TRV:CPR5 plants were much bigger than those of TRV:00 plants, indicating that silencing GhCPR5 made cotton less resistant to V. dahliae and B. cinerea. In addition, the JAZ1 expression levels in TRV:CPR5 plants were significantly higher than in TRV:00 plants, whereas the PR3, PR4, and PR5 expression levels were markedly lower in TRV:CPR5 plants.【Conclusion】GhCPR5 positively regulates cotton disease resistance; the downregulated expression of GhCPR5 significantly reduced cotton resistance to V. dahliae and B. cinerea.}
}