Dual Transcriptional Control of amfTSBA , Which Regulates the Onset of Cellular Differentiation in Streptomyces griseus

Abstract

The amf gene cluster encodes a probable secretion system for a peptidic morphogen, AmfS, which induces aerial mycelium formation in Streptomyces griseus. Here we examined the transcriptional control mechanism for the promoter preceding amfT (PamfT) directing the transcription of the amfTSBA operon. High-resolution S1 analysis mapped a transcriptional start point at 31 nucleotides upstream of the translational start codon of amfT. Low-resolution analysis showed that PamfT is developmentally regulated in the wild type and completely abolished in an amfR mutant. The ؊35 region of PamfT contained the consensus sequence for the binding of BldD, a pleiotropic negative regulator for morphological and physiological development in Streptomyces coelicolor A3(2). The cloned bldD locus of S. griseus showed high sequence similarity to the S. coelicolor counterpart. Transcription of bldD occurred constitutively in both the wild type and an A-factor-deficient mutant of S. griseus, which suggests that the regulatory role of BldD is independent of A-factor. The gel retardation assay revealed that purified BldD and AmfR recombinant proteins specifically bind PamfT. Overproduction of BldD in the wild-type cell conferred a bald phenotype (defective in aerial growth and streptomycin production) and caused marked repression of PamfT activity. An amfT-depleted mutant also showed a bald phenotype but PamfT activity was not affected. Both the bldD-overproducing wild-type strain and the amfT mutant were unable to induce aerial growth of an amfS mutant in a cross-feeding assay, which indicates that these strains are defective in the production of an active AmfS peptide. The results overall suggests that two independent regulators, AmfR and BldD, control PamfT activity via direct binding to determine the transcriptional level of the amf operon responsible for the production and secretion of AmfS peptide, which induces the erection of aerial hyphae in S. griseus.

The Pathway-Specific Regulator ClaR of Streptomyces clavuligerus Has a Global Effect on the Expression of Genes for Secondary Metabolism and Differentiation

“…A similar phenomenon occurs in S. coelicolor when the RamS peptide is added to solid cultures of a ramS mutant (52). The observed effect on amfT appears to be characteristic of ClaR and not of AdpA or BldD, which have been described to control amfT expression in S. griseus (44,53), since expression of the gene SCLAV_0719 (bldD) was not affected in S. clavuligerus ⌬claR::aac and SCLAV_1957 (adpA) was upregulated (3.7-fold) only at 60 h of culture.…”

Genes essential for morphological development and antibiotic production in Streptomyces coelicolor are targets of BldD during vegetative growth

“…For example, in contrast to S. griseus , S. coelicolor BldH does not depend on the bacterial hormone A‐factor for its expression to become derepressed (Takano et al ., 2003). S. griseus contains an orthologue of BldD (Ohnishi et al ., 2008), but, thus far, only one target gene has been identified (Ueda et al ., 2005). In this organism, BldD targets ramCSAB , a gene cluster required for the formation of AmfS, which acts as an extracellular surfactant peptide that stimulates aerial growth.…”

“…In this organism, BldD targets ramCSAB , a gene cluster required for the formation of AmfS, which acts as an extracellular surfactant peptide that stimulates aerial growth. In contrast, the orthologous system in S. coelicolor , ramCSAB, encoding the SapB surfactant peptide, is probably not targeted by BldD (Ueda et al ., 2005). We did not identify ramCSAB by ChIP‐chip with BldD, and sequence inspection of its regulatory region revealed no apparent BldD box.…”

The evolution of development inStreptomycesanalysed by genome comparisons

“…A second important morphogenetic endpoint of the cascade, that seems to bypass bldN and bldM and the production of chaplins, is the five‐gene ramCSABR cluster (= amfTSBAR in S. griseus ) (Chater & Horinouchi, 2003; Ueda et al , 2005; Willey et al , 2006). [There is a paradox in the literature here, since it was reported that a bldM mutant behaved the same as a bldD mutant in the extracellular complementation cascade (Molle & Buttner, 2000); but these experiments may be complicated by the fact that some of the mutants used to define the cascade have different genetic backgrounds.]…”