Fuente: PubMed "microbial biotechnology" Bioresour Technol. 2026 May 29:135024. doi: 10.1016/j.biortech.2026.135024. Online ahead of print.ABSTRACTSpinosad, a potent insecticidal secondary metabolite, is biosynthesized with acetyl-CoA as the key precursor. Triacylglycerols (TAGs) can supply acetyl-CoA via β-oxidation, but the regulatory link between TAG metabolism and spinosad biosynthesis in Streptomyces albus remains unclear. Here, we identified xnr_rs25305 as a key TAG hydrolase gene that positively regulates spinosad biosynthesis through transcriptomic analysis and functional validation. Overexpression of xnr_rs25305 increased spinosyn A titer by 2.1-fold and reduced intracellular TAG content by 70%, while its deletion decreased spinosyn A titer by 30%. Mechanistically, xnr_rs25305 mediates TAG hydrolysis to release free fatty acids (FFAs), which are further converted to acetyl-CoA, thereby maintaining acetyl-CoA homeostasis for spinosad biosynthesis. To augment TAG reserves, we heterologously expressed sco0958 (encoding diacylglycerol acyltransferase) in S. albus, resulting in 2.5-2.8-fold higher TAG levels during early fermentation. To coordinate TAG biosynthesis and hydrolysis temporally, we developed a novel cumate-responsive Cum-CI NOT logic gate (optimized to 5.6-fold repression efficiency) and integrated it with a cumate-inducible YES gate to construct a synthetic genetic circuit. This circuit simultaneously represses sco0958 (TAG biosynthesis) and activates xnr_rs25305 together with sco6196 (encoding acyl-CoA synthetase, accelerating FFA activation) in a cumate concentration-dependent manner. The engineered strain harboring this circuit achieved a 3.5-fold increase in spinosad titer compared to the unregulated control strain when induced with 50 μM cumate at 72 h post-inoculation. This study establishes the critical role of TAG metabolism in secondary metabolite biosynthesis and provides a precision engineering strategy by orchestrating metabolic fluxes via synthetic genetic circuits in Streptomyces.PMID:42217808 | DOI:10.1016/j.biortech.2026.135024