amfR, an essential gene for aerial mycelium formation, is a member of the AdpA regulon in the A‐factor regulatory cascade in Streptomyces griseus

Abstract

SummaryIn Streptomyces griseus , A-factor (2-isocapryloyl-3 Rhydroxymethyl-g g g g -butyrolactone) acts as a chemical signalling molecule that triggers morphological differentiation and secondary metabolism. A transcriptional activator, AdpA, in the A-factor regulatory cascade switches on a number of genes required for both processes, thus forming an AdpA regulon. amfR encoding a regulatory protein similar to response regulators of bacterial two-component regulatory systems and essential for aerial mycelium formation was found to be a member of the AdpA regulon. AdpA bound two sites at nucleotide positions approximately ----200 (site 1) and ----60 (site 2), with respect to the major transcriptional start point of amfR , and accelerated the transcription of amfR by assisting RNA polymerase in forming an open complex at an appropriate region including the transcriptional start point. Site 2 contributed more to the transcriptional activation of amfR by AdpA than site 1, although AdpA showed a much lower affinity to site 2 than to site 1. The amfR transcription enhanced by AdpA subsequently ceased at day 2 when aerial hyphae began to be formed in the wild-type strain, whereas in an adsA null mutant amfR was continuously transcribed even until day 3. This implied that amfR was repressed growth dependently by a gene product under the control of s -AdsA. Transcription of the promoter upstream of amfT depended on amfR , which is consistent with the idea that AmfR serves as an activator for amfTSBA in the amf operon. The observations that the amfR gene contains a TTA codon, a potential target for bldA -mediated regulation, and a conserved Asp-54 residue, which might be phosphorylated by a sensor kinase, suggest that the amf operon is under transcriptional, translational and post-translational control systems.

The TetR Family of Transcriptional Repressors

“…The AdpA protein in turn controls the expression of other genes. These genes include strR, which serves as a pathway-specific transcriptional activator for streptomycin biosynthetic genes (278); an open reading frame encoding a probable pathwayspecific regulator for a polyketide compound (441); adsA, which encodes an extracytoplasmic function sigma factor of RNA polymerase essential for aerial mycelium formation (438); sgmA, which encodes a metalloendopeptidase probably involved in apoptosis of substrate hyphae during aerial mycelium development (174); ssgA, which encodes a small acidic protein essential for spore septum formation (437); amfR, essential for aerial hyphae formation (398,440); and the sprT and sprU genes, which encode trypsin-like proteases (173).…”

DNA microarray analysis of global gene regulation by A-factor in Streptomyces griseus

“…The AraC/ XylS family transcriptional regulator AdpA then activates transcription of many genes that are required for secondary metabolism and morphological differentiation, forming an AdpA regulon (Ohnishi et al, 2005). Members of the AdpA regulon identified to date are strR, the pathway-specific transcriptional activator for streptomycin biosynthesis (Ohnishi et al, 1999;Tomono et al, 2005a); an ORF (SGR6071) encoding a probable pathway-specific transcriptional activator for biosynthesis of a polyketide compound (Yamazaki et al, 2004); adsA, encoding an extracytoplasmic function sigma factor of RNA polymerase essential for aerial mycelium formation ; amfR, encoding a transcriptional activator for the amfTSBA operon responsible for production of a surfactant lantibiotic-like peptide SapB (AmfS) (Ueda et al, 2002(Ueda et al, , 2005, which is essential for aerial mycelium formation (Yamazaki et al, 2003b); ssgA, encoding a small acidic protein essential for spore septum formation (Yamazaki et al, 2003a); sgmA, encoding a secreted metalloendopeptidase probably involved in apoptosis of substrate mycelium during aerial mycelium development (Kato et al, 2002); five different genes (sprA, sprB, sprD, sprT and sprU) encoding secreted serine proteases (Kato et al, 2005a;Tomono et al, 2005b); sgiA, encoding a Streptomyces subtilisin inhibitor (SSI) family inhibitor (Hirano et al, 2006); a gene of unknown function (SGR4618, formerly named orfA-AdBS3) (Yamazaki et al, 2004); and an operon of unknown function orf1-2-3-4 of AdBS2 (SGR6559-SGR6556) (Yamazaki et al, 2004). Thus, we have identified 14 AdpA target promoters to date.…”

AdpA, a Central Transcriptional Regulator in the A-Factor Regulatory Cascade That Leads to Morphological Development and Secondary Metabolism inStreptomyces griseus

“…22,23) Among the DNA fragments, we found adsA that encodes an extracytoplasmic function (ECF) sigma factor of RNA polymerase essential for aerial mycelium formation, 22) sgmA that encodes a metalloendopeptidase that is probably involved in apoptosis of substrate hyphae during aerial mycelium development, 24) ssgA that encodes a small acidic protein essential for spore septum formation, 25) and an open reading frame that encodes a probable pathway-specific transcriptional regulator for the biosynthesis of a polyketide compound. 20) In addition to these genes, we also found the following genes as targets of AdpA through genetic studies: amfR that encodes a response regulator-like protein essential for activation of the amf operon, 26) sprT and sprU, both of which encode a trypsin-type protease, 27) and sprA, sprB, and sprD, all of which encode a chymotrypsin-type protease (unpublished results). There must be many other genes controlled by AdpA.…”

“…For transcriptional activation, some genes require simultaneous binding of a dimer of AdpA to multiple sites. Despite the differences in the binding position and the number of binding sites, AdpA recruits RNA polymerase to the specific promoter region of the target genes and facilitates the isomerization of the RNA polymerase-DNA complex into an open complex for transcriptional initiation, as determined by potassium permanganate footprinting for amfR, 26) adsA, 20) and ssgA. 20) Recruitment of RNA polymerase to the promoter of a target gene is a typical mechanism for the transcriptional activation of general activators.…”