show Abstracthide AbstractStreptomyces possess the genetic potential to produce diverse clinically important secondary metabolites. Many biosynthetic gene clusters (BGCs) is often activated by interaction with . One of the approaches to activate the silent BGCs is microbial co-culture which mimicking the natural ecological habitat of the microbes. Co-culture of Streptomyces coelicolor with Myxococcus xanthus which prey on a wide variety of microbes induces overproduction of actinorhodin by S. coelicolor. Here, we performed transcriptome analysis of M. xanthus and S. coelicolor during co-culture to elucidate the underlying mechanism how BGC for actinorhodin biosynthesis is activated. During the co-culture, S. coelicolor activated the actinorhodin biosynthesis and branched amino acid degradation to provide precursor, acetyl-CoA. Simultaneously, M. xanthus enhanced the production of siderophore myxochelin and the iron acquisition system. Myxochelin itself and chemical compound produced by M. xanthus had no effect, but the active chemical communication between S. coelicolor and M. xanthus was required for triggering actinorhodin overproduction of S. coelicolor. Iron level measurement and iron-rich co-culture revealed that M. xanthus mediated iron-depleted environment stimulated actinorhodin overproduction of S. coelicolor. This study utilized transcriptome analysis to determine the veiled mechanism, which stimulates the production of the secondary metabolite cluster during the co-culture.