Regulation Of Phenylpropanoids Biosynthesis By MYB,bHLH And WD40 Transcription Factors In Solanaceae

Phenylpropanoids are secondary metabolisms based on aromatic rings as skeleton which are from phenylalanine catalyzed by a series of enzymes.They can not only protect plants from biotic and abiotic stress.For example,they can raise resistance to drought,cold,salt,plant diseases and insect pests.Our research isolated several MYB,bHLH and WD40 transcription factors from tomato and Solanum nigrum.Studying the effect of those transcription factors on regulating phenylpropanoids biosynthesis provides a good basis for breeding tomatoes with high phenylpropanoids contents and high resistance to plant diseases and insect pests.We firstly compared phenylpropanoids contents in AtMYB12/Ros1/Del-expressing(ADR),Ros1/Del-expressing(DR)and AtMYB12-expressing(A)tomatoes.Contents of phenylpropanoids including flavonols,anthocyanins and caffeoyl quinic acids in the former are higher than that of the two latter.We designed a comparing method of transcriptome sequencing between three transgenic tomatoes.5(4 up-regulation and 1 down-regulation),9(6 up-regulation and 3 down-regulation)and 1(down-regulation)candidate genes related to flavonols,anthocyanins and caffeoyl quinic acids biosynthesis were obtained,respectively.Then we verified candidate genes regulating phenylpropanoids biosynthesis by determining expression levels in different development stages of three transgenic tomatoes,which is consistent with the result of transcriptome sequencing.Four candidate transcription factors could interact with AtMYB12 and Rosea1,which ulteriorly verified their effects on phenylpropanoids biosynthesis.The next purpose of our research is mainly exploring effects of candidate transcription factors on regulating phenylpropanoids biosynthesis after they were transformed into tomatoes and the effect of phenylpropanoids on resistance of transgenic tomatoes to plant diseases and insect pests.Solanum nigrum also belongs to Solanaceae plants.In our study,we determined the anthocyanins content was up to 3.95 mg/g and there was a positive correlation between anthocyanins content and antioxidant ability with r=0.93.So we cloned an R2R3-MYB gene named SnMYB by RACR-PCR in the circumstance of unknowing the genomic sequences of Solanum nigrum.The protein sequence of SnMYB has three anthocyanin-related motifs and it can successfully induce anthocyanins production when transiently expressed in tobacco leaves(1.03 mg/g).Those evidences comfirmed that SnMYB can positivly regulate anthocyanins biosynthesis.Our study provides good basis for exploring more anthocyanins as natrual antimicrobial to prevent plant diseases and utilizing SnMYB to breed transgenic tomato with high anthocyanins content.Then four fruit specific promoters E8,E4,2A12 and PG drove At MYB12 gene expressed in the same tomato variety.The four promoters could drive At MYB12 gene in a higher expression level with 21.55,5.43,7.78,6.45 fold compared to wild tomatoes.Correspondingly,the four promoters drove AtMYB12 increasing total contents of phenylpropanoids with 15.51,6.41,5.44,4.29 fold,which indicated that different fruit promoters had different effects on regulating phenylpropanoids biosynthesis of AtMYB12.This result will help our candidate transcription factor genes to express in tomatoes with the maximum efficiency.Finally,our research expressed SlMYB12 under the E8 promoter in three different tomato varieties.In three wild tomato varieties,we found that there was positive correlation between flavonols contents and expression level of SlMYB12 gene with r=0.96.A well positive correlation was also between flavonols contents and antioxidant activity with r=0.99 in three wild tomato varieties.In three SlMYB12-expressing tomatoes,there also was positive correlation between rutin and kaempferol contents and expression level of SlMYB12 gene with r=0.98 and r=0.95.Those results indicated that SlMYB12 could not only regulate flavonols biosynthsis but also be used as an endogenous marker gene in chosing good tomato variety to maximum the effect on phenylpropanoids regulation of candidate genes.