Engineering the TetR-family transcriptional regulator XNR_0706 to enhance heterologous spinosad production in Streptomyces albus B4 chassis

The TetR family of regulators are an important group of transcription regulators that regulate diverse cellular processes in prokaryotes. In this study, we found that XNR_0706, a TetR family regulator, controlled the expression of <i>XNR_0345</i>, <i>XNR_0454</i>, <i>XNR_0513</i> and <i>XNR_1438</i> putatively involved in fatty acid β-oxidation by interacting with the promoter regions in <i>Streptomyces albus</i> B4. The transcription level of these four genes was downregulated in <i>XNR_0706</i> deletion strain (ΔXNR_0706) and restored by <i>XNR_0706</i> complementation in Δ0706/pIB-<i>0706</i>, demonstrating that XNR_0706 was a positive transcriptional regulator of the genes. With toxic long-chain fatty acids addition in TSB media, deletion of <i>XNR_0706</i> caused significantly poor growth, whereas <i>XNR_0706</i> complementation increased the utilization of additional fatty acids, resulting in restored growth. Fatty acid β-oxidation is one source of acetyl- and malonyl-CoA precursors for polyketides biosynthesis in actinobacteria. Overexpression of <i>XNR_0706</i> in B4/spnNEW, a spinosad heterologous expression strain derived from <i>S. albus</i> B4, increased spinosad yield by 20.6 %. Additionally, supplement of 0.3 g/L fatty acids resulted in a further 42.4 % increase in spinosad yield. Our study reveals a regulatory mechanism in long-chain fatty acids metabolism in <i>S. albus</i> and these insights into the molecular regulation of β-oxidation by XNR_0706 are instrumental for increasing secondary metabolites in actinobacteria.