'红满堂'苹果MbbZIP43基因的克隆与功能研究OA北大核心CSTPCD

[Objective]To investigate the regulatory role of basic leucine zipper(bZIP)transcription factors in anthocyanin metabolism in'Hongmantang'apple.[Method]We cloned a bZIP gene from'Hongmantang'red-flesh apple,analyzed it and its sequence characteristics,studied the subcellular localization of its encoded protein using onion epidermis cell transient overexpression,and investigated its expression pattern in'Hongmantang'red-flesh apple fruits at different ripening stages using quantitative real time PCR(RT-qPCR).Furthermore,we explored its regulatory roles in the expressions of genes related to anthocyanin accumulation and biosynthesis based on the transient overexpression experiments in tobacco leaves,apple leaves and fruits.[Result]This bZIP gene was intron-less and shared the highest similarity with MdbZIP43(XP_008393381.1),thus was named as MbbZIP43.Its coding sequence was 522 bp,which encoded a 173 aa hydrophilic nucleus-localized protein with conserved bZIP_plant_GBF1 domain.RT-qPCR analysis revealed that MbbZIP43 showed a'rise-fall'expression change pattern in'Hongmantang'fruits during ripening,with the highest expression in the fruits at 11 weeks post flowering.Correlation analysis results showed that MbbZIP43 expression was positively correlated with the total flavonoids and anthocyanin contents in'Hongmantang'fruits,indicating that it might play regulatory roles in flavonoid biosynthesis.Consistently,transient overexpression of MbbZIP43 improved significantly the anthocyanin accumulations by 17.42%,25.66%and 48.99%in tobacco leaves,apple leaves and apple fruits,respectively.Moreover,RT-qPCR analysis results showed that,compared to the empty vector control,the expressions of anthocyanin biosynthesis related structural genes(CHI,F3'H,DFR and UFGT)in the apple leaves overexpressing MbbZIP43 were up-regulated by 2.27-,1.84-,2.39-and 2.89-fold,respectively.And their expressions in the peels of apple fruits overexpressing MbbZIP43 were up-regulated by 1.79-,1.70-,1.35-and 1.51-fold,respectively.These results indicated that the gene's transient overexpression may improve anthocyanin accumulation by activating the expressions of the anthocyanin biosynthesis related structural genes.[Conclusion]MbbZIP43 can promote anthocyanins accumulation by activating the expression of anthocyanin biosynthesis related structural genes,including CHI,UFGT and so on.