Mechanism Of NAC Transcription Factor MdNAC52 Regulating Biosynthesis Of Apple Anthocyanins And Proanthocyanidins

Apple is one of the most important fruit trees in the world.It is very important to improve the quality of apple fruit.Among them,the color of the fruit is a very important agronomic trait,and anthocyanins are the main pigments that determine the coloring of apples.There are many factors affecting its biosynthesis,such as light,hormones,and temperature.Proanthocyanidins are flavonoids whose content can affect the nutritional quality of fruits.Anthocyanins and proanthocyanidins play important roles in plant growth and development.Although previous studies have identified many transcription factors related to the anthocyanin and proanthocyanidin pathways,the regulatory mechanism of these pathways remains to be further studied.Sequencing the transcriptome of different budding apple cultivars in the early stage,and analyzing the results,I found a NAC transcription factor MdNAC52,whose gene transcript level showed differences in red apples with different degrees.So we speculate that MdNAC52 is a regulator related to anthocyanin metabolism.In this study,I used 'Yanfu 3' as a test material and carried out research on gene function verification by cloning MdNAC52 transcription factor.The main results are as follows:1.In 'Yanfu 3' apple,the anthocyanins of the peel gradually accumulated from 2 to 12 days after picking,and reached a peak at 12 days;during this coloring period,the proanthocyanidin content decreased.RT-PCR was used to detect the expression pattern of MdNAC52,and it was found that the expression level reached a peak on the 8th day after bag removal,and then decreased.2.MdNAC52 was overexpressed in apple ‘orin' callus,and it was found that apple callus would turn red,this process not only promoted anthocyanin synthesis in callus but also promoted structural genes such as MdANS and MdUFGT in the anthocyanin synthesis pathway.3.Yeast one-hybrid and in vitro interaction experiments EMSA have verified that MdNAC52 can specifically bind MdMYB9 and MdMYB11,thereby promoting anthocyanin synthesis.In plants,ChIP experiments have verified that MdNAC52 interacts with MdMYB9 and MdMYB11.4.MdNAC52 expression was induced by light and the expression pattern was positively correlated with MdHY5.The cis-elements on the MdNAC52 promoter were analyzed and found to contain G-box.Yeast one-hybrid,EMSA,and ChIP experiments were performed to verify that MdNAC52 is downstream of MdHY5 and induced by light.5.The proanthocyanidin content in overexpression ‘orin' callus was significantly increased,and so was the expression of MdLAR.Yeast one-hybrid,in vitro interaction experiments EMSA,and in vivo ChIP experiments verified that MdNAC52 interacts with MdLAR,and then promotes the accumulation of proanthocyanidins in apple callus.This study demonstrated that MdNAC52 binds to MdMYB9 and MdMYB11 promoters and promotes the accumulation of anthocyanins and proanthocyanidins.And it directly binds to MdLAR to regulate the synthesis and accumulation of proanthocyanidins.This research provides new insights into the role of NAC transcription factors in regulating apple anthocyanins and proanthocyanidins accumulation.