Functional Characterization Of GhCNGC13 And GhCNGC32 In Upland Cotton

Upland cotton(Gossypium hirsutum L.)is one of the most important economic crops in China.For a long time,however,cotton production is susceptible to serious impacts of various biotic and abiotic stresses,which has become a prominent problem that limits the increase of cotton production and improvement of fiber quality.It is of great importance to gain in-depth understanding about the basic theoretical knowledge of cotton biology,discover and utilize key regulatory genes for the development of new cotton varieties with higher yield,stress tolerance and disease resistance which warrants the continuous development of cotton industry in our country.Cyclic nucleotide gated channels(CNGCs)are a family of non-selective cation channels that are widely present in different species.Recent research in the past years has shown that members of this family are involved in many important biological processes such as plant growth and development as well as environmental stress responses in Arabidopsis.However,there is still a lack of understanding about the regulatory roles of CNGCs toward agronomic traits in crops.Up to now,little is known about CNGCs in cotton.In this work,we conducted a preliminary study of CNGCs genes in upland cotton.The major results are as follows:1.A total of 40 CNGCs were identified in upland cotton genome through bioinformatics analysis,named Gh CNGC1-40,respectively.These genes were localized on 19 different chromosomes.Phylogenetic analysis based on their protein sequences showed that Gh CNGC1-40 were clearly divided into groups I,II,III and IV,of which group IV comprised two subgroups IV-A and IV-B.2.Gene expression patterns of Gh CNGCs were examined using quantitative realtime PCR(q RT-PCR),showing that most Gh CNGCs expressed in different organs and/or tissues.Moreover,these genes showed differential expression responses under various treatment of salt stress(Na Cl),hormones(ABA and Me JA)and fungal infection of Verticillium dahliae(Vd991).3.Virus-induced gene silencing(VIGS)was performed with Gh CNGC13 and Gh CNGC32 in cotton plants,and the results showed that transcriptional reduction of Gh CNGC13 and Gh CNGC32 significantly suppressed plant height,root and leaf development,resulted in the accumulation of reactive oxygen species and appearance of necrotic spots in leaves,as well as significant changes in the levels of various endogenous hormones.In addition,the silenced plants displayed significantly reduced resistance to Verticillium wilt.These results indicate that Gh CNGC13 and Gh CNGC32 play critical roles in the growth and development of cotton plants and Verticillium wilt resistance.4.Transcriptome comparative analysis between Gh CNGC13 and Gh CNGC32-silenced plants and control plants was carried out using RNA-Seq technique,and 3546up-regulated and 1289 down-regulated differentially expressed genes were identified.GO function enrichment analysis showed that the significantly up-regulated genes were mainly related to biological processes such as protein phosphorylation and metabolism,while the significantly down-regulated genes were mainly related to photosynthesis and carbon utilization.KEGG enrichment analysis showed that these differentially expressed genes played crucial roles in the pathways of plant pathogen interaction,photosynthesis and MAPK signaling.These results support the phenotypic effects of the silenced plants,which laid a solid foundation for further in-depth study.