Studies On The Expression And Function Of Transcription Factors Related To Abiotic Stresses In Haematococcus Pluvialis

Haematococcus pluvialis Flotow is considered to be an excellent source of natural astaxanthin becase it can accumulate large amounts of astaxanthin under stress conditions.At present,the research on H.pluvialis mainly focuses on promoting the production and its extraction technology.However,there are relatively few studies on the regulation of astaxanthin synthesis in H.pluvialis,result in being unclear to its mechanism.The purpose of this study was to investigate the regulatory role of transcription factors in the accumulation of astaxanthin in H.pluvialis under environmental stress.High-throughput sequencing was carried out on H.pluvialis 192.80,further to analyze the expression trends of differentially expressed genes(DEGs)and screen differentially expressed transcription factors.Full-length sequences of transcription factors were cloned,and their structures and functions were analyzed,in order to clarify the relationship between these transcription factors and the key enzyme genes of astaxanthin synthesis.The specific research content and results are as follows.(1)High-throughput sequencing was performed after treated H.pluvialis 192.80 with high light intensity and sodium acetate.The expression trend analysis,GO enrichment analysis and Pathway enrichment analysis was proceeded.332 differentially expressed transcription factors,belonged to 28 families respectively,were predicted.They showed a variety of expression patters to adapt enviromenatal changes.At the same time,H.pluvialis cells accumulated astaxanthin.The synthesis of secondary metabolites in H.pluvialis was vigorous,so as to protect the cells from stress-promoting damage.This indicates that there might be a certain relationship between transcription factors and the synthesis of secondary metabolites.(2)It was found that the astaxanthin production in nitrogen deficiency group was the highest among different stresses including high-intensity light,sodium acetate,salicylic acid,and nitrogen deficiency.Based on the third-generation transcriptome data,Haematococcus pluvialis 8 transcription factors from AP2,b HLH,b ZIP,ERF,G2-like,HSF,Whirly and WRKY families were cloned.All of them contained the conserved domains of TF families.Further analysis of the transcription level revealed that transcription factors might play an important role under different stress conditions.(3)The promoter sequences of BKT1,BKT2,Crt R-b1,and Crt R-b2 were found after mining genome data of H.pluvialis.The analysisi of transcription factor binding sites in these promoters showed that transcription factors such as ERF and Nin-like might play an important role in the transcriptional regulation of BKT1,BKT2,Crt R-b1 and Crt R-b2.(4)Nine ERF transcription factors,which contained AP2/ERF domains and showed high similarity.The results of subcellular localization prediction showed that most of them were localized to the nucleus.The Hp ERF transcription factors had homology with Arabidopsis thaliana.(5)The expression of 9 Hp ERF transcription factors and BKT1,BKT2,Crt R-b1 and Crt R-b2 genes were up-regulated in different degrees,under different stress conditions.The results of correlation analysis showed that the expression of 9 Hp ERF transcription factors and 4 astaxanthin synthesis key enzyme genes were high in H.pluvialis.In summary,we can speculate that under high-intensity light,nitrogen deficiency and other stress conditions,the expression of transcription factors such as Hp ERF are up-regulated to respond to different environmental stimuli.They might bind the promoters of key enzyme genes related with astaxanthin synthesis,such as BKT1,BKT2,Crt R-b1 and Crt R-b2,to fulfill regulating the astaxanthin synthesis.Hence,the above research laied the foundation for us to clarify the transcriptional regulation of key enzyme genes for astaxanthin synthesis guided by transcription factors.