Mechanisms Of CsMYB6 And CsTS1 In Regulating Fruit Spine And Tubercule Development In Cucumber

The cucumber(Cucumis sativus L.)warty fruits have both spines and tubercules on their surface The warty(Wty)trait has an important impact on the commercial value of cucumber fruit.Compared with large-warty(L-Wty)cucumber fruits,small-warty(S-Wty)fruits possess a smoother appearance and are more easily cleaned,packed,stored,and transported.There is therefore great interest in identifying the molecular mechanism that controls the formation of cucumber fruit spines and tubercules.However,the molecular mechanism of spines and tubercules development is not well understood.1.Using an analysis transcriptome profiling data between WT and glabrous mutants,CsMYB6 was chosen as a candidate gene which affect the formation of cucumber fruit spine.we report functional characterization of the cucumber CsMYB6 gene,which encodes a MIXTA-like MYB transcription factor that plays an important role in regulating fruit trichome development.Spatial-temporal expression analyses revealed high-level expression of CsMYB6 in the epidermis of cucumber ovaries during fruit spine initiation,which was similar to the expression of CsTRY,a homolog of the Arabidopsis TRY gene that also plays a key role in trichome development.Overexpression of CsMYB6 and Cs TRY in cucumber and Arabidopsis revealed that CsMYB6 and CsTRY act as negative regulators of trichome initiation in both species,and that CsMYB6 acted upstream of CsTRY in this process.CsMYB6 was found to bind to the three MYB binding sites inside the promoter region of CsTRY,and protein-protein interaction assays suggested that CsTRY also directly interacted with CsMYB6 protein.2.The results of the genetic analysis showed that single dominant gene,TS1(Tubercule size gene ?),controls the L-Wty fruit trait.CsTS1 was identified by map-based cloning and RNA-seq methods,and was found to encode an oleosin protein.High CsTS1 expression levels were closely correlated with increased fruit tubercule size among 44 different cucumber lines.Transgenic complementation and RNAi-mediated gene silencing of CsTSl in transgenic cucumber plants demonstrated that CsTS1 positively regulates the development of tubercules.Spatial-temporal expression analyses revealed CsTSl is highly expressed in the peel at fruit tubercule forming and enlargement stage.Allelic variation was identified in the promoter region of CsTSl,resulting in low expression of CsTSl in all 22 different small-warty or non-warty cucumber lines.Molecular and genetic analyses show that CsTS1 acts downstream of CsTu,a key tubercule formation factor,in regulating the development of fruit tubercules.Using yeast one-hybrid assay and dual luciferase reporter assay,we determined that CsTu directly binds to the CsTS1 promoter and promotes its expression.