The Function Analysis Of Cotton HD1 In The Regulation Of Anthocyanin Synthesis And The Development Of Stem Trichom

GaHD1 gene is a member of the Homeodomain-leucine zipper IV family,which plays an important role in cotton fiber development.Previous studies found that in addition to affecting the cotton fiber phenotype,GaHD1 over-expressed Arabidopsis thaliana also exhibited purple leaves and weak root architecture,indicating that the function of the HD1 gene may have multiple effects,for example,is involved in the regulation of anthocyanin biosynthesis.In the study of the regulation of anthocyanin synthesis and related gene's function,the members of the HDZIP-IV gene family in which GaHD1 is located have not been well studied.Therefore,this study firstly analyzed the anthocyanin content and other phenotypes of transgenic cotton and arabidopsis to preliminarily identify the role of GaHD1 gene in anthocyanin synthesis;By comparison of transcriptome data generated by RNA-seq,the differentially expressed genes of transgenic Arabidopsis thaliana with overexpressed GaHD1 were analyzed and verified by RT-PCR to identify GaHD1 downstream genes regulating anthocyanin synthesis;Based on the obtaining differentially expressed genes,further bioinformatics analysis was performed to perform cis-acting element enrichment analysis on their promoters.The main results are as follows:1.Phenotypic analysis of Arabidopsis overexpressing GaHD1 showed that transgenic Arabidopsis root system are abnormally developed compared to the wild type,but its anthocyanin content was significantly higher than that of wild type.The results of PEG-treated drought stress experiments showed that the over-expressing GaHD1 Arabidopsis thaliana can rapidly accumulate the anthocyanin,and its anthocyanin accumulation rate was significantly higher than that of the wild type.2.The Anthocyanin content,stem trichome and fiber quality of HD1 over-expression cotton were measured and analyzed respectively.We found that Anthocyanin content,trichome density,and fiber length are significantly higher than the wide-type cotton.3.Transcriptome analysis of the GaHD1 over-expression Arabidopsis' s leaves has identified 1335 up-regulated genes and 2308 down-regulated genes.Among the up-regulated genes,18 members of the MYB family are found to be related to anthocyanin synthesis.Gene Ontology(GO)and KEGG pathway enrichment analysis were also performed for the up-regulated genes.The results indicate that the most enriched pathways are “responded to stimuli” and “responded to stress” which contain a large number of MYB transcription factors;The anthocyanin synthesis pathway is the pathway with the highest percentage of enriched genes,which also indicates that the HD1 gene may increase the expression of critical genes that regulate anthocyanin synthesis,and thus increase the anthocyanin content in Arabidopsis.4.In Arabidopsis,sequence analysis of the selected transcription factors(TT4,TT8,MYB11)found that the three transcription factor promoter sequences all contained the cis-acting element L1-box bound by the HD1 gene.The results of real-time quantitative PCR experiments showed that the expression levels of these transcription factors in the over-expressed GaHD1 Arabidopsis were significantly higher than those of the wild type,indicating that the HD1 gene may regulate the related transcription factors in the anthocyanin biosynthesis pathway to generate anthocyanin.