R3 MYB-like Transcription Factor PtrRML1 Regulates Epidermal Cell Fate Determination And Anthocyanin Biosynthesis In Arabidopsis

MYB transcription factors are one of the biggest transcription factor families in plants.MYB proteins are characterized by a highly conserved DNA-binding domain: the MYB domain.This domain generally consists of up to four imperfect amino acid sequence repeats(R)of about 52 amino acids.Depending on the number of adjacent repeats,MYB transcription factors can be divided into 4R-MYB(R1R2R2R1/2 MYB),3R-MYB(R1R2R3 MYB),2R-MYB(R2R3 MYB)and 1R-MYB(R1/2 MYB,R3 MYB,MYB-related and others).Single-repeat R3 MYB transcription factors(R3 MYBs)are small proteins(~100 amino acids)that largely consists of a single MYB DNA-binding repeat,and are best characterized for their regulatory roles in epidermal cell fate determination.R3 MYBs regulate epidermal cell fate determination in Arabidopsis through a lateral inhibition mechanism.R3 MYBs compete with R2R3 MYBs for binding bHLH transcription factors,thus blocking the formation of the MYB-bHLH-WD40(MBW)activator complexes,thereby inhibiting the expression of GL2,leading to the inhibition of trichome formation and promotion of root hair formation.The MBW activator complexes that regulate epidermal cell fate determination in Arabidopsis are formed by three different types of transcription factors,including R2R3 MYB transcription factors GLABRA1(GL1),WEREWOLF(WER),or AtMYB23,bHLH transcription factors GL3,or ENHANCER OF GLABRA3(EGL3),and the WD40-repeat protein TRANSPARENT TESTA GLABA1(TTG1).Similar MBW complexes can also regulate anthocyanin biosynthesis in Arabidopsis.Anthocyanin biosynthesis are catalyzed by a series of enzymes,genes encoding these enzymes are divided into early and late biosynthesis genes.The early biosynthesis genes are regulated by R2R3 MYB transcription factors,and the late biosynthesis genes are regulated by MBW complexes.The MBW activator complexes involve in the regulation anthocyanin biosynthesis are formed by an R2R3 MYB transcription factors PRODUCTION OF ANTHOCYANIN PIGMENT 1(PAP1)?PAP2?MYB113 or MYB114,a bHLH transcription factor TRANSPARENT TESTA 8(TT8),GL3 or EGL3,and the WD40-repeat protein TTG1.We report here the identification and functional characterization of a poplar R3 MYB-like protein,Populus trichocarpa R3 MYB-LIKE1(PtrRML1).The amino acid number of PtrRML1(199aa)is about twice of that of R3 MYBs.PtrRML1 has a typical R3 MYB repeat at its C-terminal.PtrRML1 is localized in nuclear as examined in transgenic Arabidopsis plants expressing PtrRML1-GFP.When tested in protoplasts,PtrRML1 repressed reporter gene expression,and it interacted with GL3.Expression of PtrRML1 in Arabidopsis resulted in glabrous phenotypes,increased number of root hairs,and decreased anthocyanin accumulation.In consist with these observation,the expression levels of some MBW component genes and some of their target gene involved in the regulation of epidermal cell determination and anthocyanin accumulation,including TRYPTICHON(TRY),CAPRICE(CPC),ENHANCER OF TRY AND CPC1(ETC1),GL1,GL2,DIHYDROFLAVONOL REDUCTASE(DFR),UDP-FLAVONOID 3-GLUCOSYL TRANSPARENT TESTA(UF3GT)and TT8 were reduced in the transgenic plants.