Daptomycin,a member of cyclic lipopeptides produced by Streptomyces roseosporus,is a critical antibiotic against Grampositive bacteria.Although there are many reports on the metabolite biosynthetic pathway,genetic manipulation,clinical application of daptomycin,the biosynthetic related genes are rarely characterized by now.The dptP gene shares a significant sequence similarity with membrane proteins of streptomyces multispecies,mainly membrane protein DedA.The dptP gene locates upstream of the dpt gene cluster(Figure 1).Here we constructed a series of expression plasmids to decipher the functional characterization of dptP gene of Streptomyces roseosporus(Figure 2).The deletion of dptP by a CRISPR-Cas9 system(Figure 3A)decreased the transcription levels of the biosynthetic genes including dptE,dptF,dptM,dptN,dptA,dptBC and dptD.And overexpression(Figure 3B)of dptP induced the reverse effects(Figure 4),which increased the yield of daptomycin.These results suggested that dptP plays a positive regulation role for daptomycin biosynthesis in Streptomyces roseosporus.Our discovery of dptP-mediated metabolism regulation helps to rational design and improve daptomycin biosynthesis.