Inactivation of the sco2730/2731 copper chaperone-transporter system in Streptomyces coelicolor and its orthologs in Streptomyces venezuelae, together with chromosomal end deletion, greatly enhances secondary metabolism.

BACKGROUND: Activating silent biosynthetic gene clusters (BGCs) in Streptomyces remains a major challenge in harnessing their vast secondary metabolic potential and requires diverse, complementary strategies, including bacterial co-cultivation, heterologous expression, chemical elicitation, modulation of gene expression, and the use of pleiotropic and pathway-specific genetic regulators, such as those influenced by cytosolic copper levels. Previous studies reported that, in S. coelicolor, disruption of the sco2730/2731 copper chaperone–transporter system (Sc-M1 mutant) markedly enhances secondary metabolism. However, this activation is only partially reproduced by antisense knockdown constructs targeting sco2730/sco2731 in S. coelicolor (Sc-M2 mutant) and other species (S. venezuelae, S. albidoflavus). This study investigates the basis of the strong activation observed in Sc-M1, with the aim of exploiting this mechanism for activating silent BGCs in Streptomyces. RESULTS: Genomic analysis revealed that, in addition to sco2730/2731 inactivation, the Sc-M1 mutant possesses a spontaneous deletion of both chromosomal ends. Construction of a sco2730 knockout mutant (Δsco2730, also affecting sco2731; Sc-M3 mutant) showed an effect on secondary metabolism comparable to that of the Sc-M2 mutant, and demonstrated that sco2730 disruption increases chromosomal-end instability. Metabolomic analyses showed that inactivation of sco2730/2731 (Sc-M3 mutant) or chromosomal-end deletion (Sc-M4 mutant) individually enhanced secondary metabolism. However, only the combination of Δsco2730 and chromosomal-end deletion (Sc-M5 mutant) approached the extensive metabolic activation observed in Sc-M1, affecting up to 60 secondary-metabolite adducts from 17 biosynthetic pathways. Similar synergistic effects were observed in S. venezuelae, where combined knockdown of the sco2730/2731 orthologues and chromosomal-end deletion strongly modulated secondary metabolism, repressing chloramphenicol production while inducing pikromycin biosynthesis, a typically silent and difficult-to-activate S. venezuelae BGC. CONCLUSIONS: Simultaneous disruption of the sco2730/31 copper chaperone–transporter system and chromosomal-end deletion synergistically enhance secondary metabolism production in S. coelicolor and S. venezuelae. This combined genetic manipulation provides a novel strategy for the challenging task of activating silent biosynthetic pathways and for potentially discovering new bioactive compounds across Streptomyces species.