amfC plays a regulatory role in aerial mycelium formation in Streptomyces griseus and is distributed widely among Streptomyces species. Disruption of the chromosomal amfC gene in Streptomyces coelicolor A3(2) severely reduced formation of aerial hyphae, indicating that amfC is important in morphological development. In addition, the disruption caused S. coelicolor A3(2) M130 to produce much less actinorhodin, and to produce undecylprodigiosin at a later stage of growth, indicating that amfC also regulates secondary metabolism. S1 nuclease mapping showed that transcription of actII-ORF4, the pathway-specific transcriptional activator in the act gene cluster, was greatly reduced in the amfC disruptants. The defect in secondary metabolite formation was suppressed or overcome by a mutation in sre-1. Consequently, an amfC-disrupted strain derived from S. coelicolor A3(2) M145, an actinorhodin-overproducing strain due to the sre-1 mutation, still produced a large amount of actinorhodin. Extra copies of amfC in strains M130 and M145 did not change spore-chain morphology or secondary metabolite formation. However, the spores in these strains remained white even after prolonged incubation. Since only spore pigmentation was affected, all known whi genes, except whiE, responsible for the polyketide spore pigment formation, were assumed to function normally. S1 nuclease mapping showed that transcription of whiEP1, one of the promoters in the whiE locus, was reduced in S. coelicolor A3(2) containing extra copies of amfC. Introducing amfC into several other Streptomyces species, such as Streptomyces lividans, Streptomyces lavendulae and Streptomyces lipmanii, also abolished spore pigment formation. An increase in the amount of AmfC appeared to disturb the maturation of spores.