A portable pathway specific transcription factor-promoter circuit enables heterologous production of fungal nonribosomal peptides

Source: PubMed Central Open Access, NCBI / U.S. National Library of Medicine

Most fungal nonribosomal peptide synthetase (NRPS) biosynthetic gene clusters remain transcriptionally silent, severely restricting the discovery of bioactive peptide metabolites. Standard constitutive promoters fail to support robust expression of large multidomain NRPS enzymes. Here, we identify a conserved subgroup of Zn()Cystranscription factors (TFs) encoded within fungal NRPS clusters and functionally characterize TdaN, which triggers transcription from its cognate promoter Pby recognizing paired CGG-type-regulatory motifs. The TdaN–Ppair is repurposed as a portable transcriptional circuit capable of activating five out of seven heterologous NRPS/NRPS-like genes in, notably the recalcitrant silent synthetase Afpes1 that previously demanded screening over 90 distinct promoters for trace product yields. Circuit-driven overexpression ofyields abundant--Trp--Pro (cWP) and seven structurally diversified cWP derivatives. This study reveals a conserved regulatory paradigm for fungal nonribosomal peptide biosynthesis and delivers a versatile transcriptional module to expand cryptic peptide chemical diversity. A native fungal PSTF-promoter circuit is repurposed as a portable transcriptional tool box to activate silent NRPS pathways, enabling heterologous production of diverse nonribosomal peptides and expanding fungal natural product diversity. toc