Identification of an A-factor-dependent promoter in the streptomycin biosynthetic gene cluster of Streptomyces griseus

Abstract

A-factor (2-isocapryloyl-3R-hydroxymethyl-gamma-butyrolactone) is a microbial hormone controlling streptomycin (Sm) production, Sm resistance and sporulation in Streptomyces griseus. In order to identify A-factor-dependent promoters in the Sm biosynthetic gene cluster, a new promoter-probe plasmid with a low copy number was constructed by using an extremely thermostable malate dehydrogenase gene as the reporter. Of the three promoters in the Sm production region that includes strR, aphD and strB, only the promoter of strR, which codes for a putative regulatory protein, was found to be directly controlled by A-factor. This was also confirmed by S1 nuclease mapping. The region essential for its A-factor-dependence was determined to be located 430-330 base pairs upstream of the transcriptional start point. Increase in the copy number of the strR promoter region did not lead to a corresponding increase in the total promoter activity, probably due to titration of a putative activator which binds to the enhancer-like region and controls the expression of the strR promoter. This putative activator is apparently distinct from the A-factor-receptor protein. The aphD gene, which encodes the major Sm resistance determinant, Sm-6-phosphotransferase, was transcribed mainly by read-through from the A-factor-dependent strR promoter; this accounts for the prompt induction of Sm resistance by A-factor.

A gene cluster involved in aerial mycelium formation in Streptomyces griseus encodes proteins similar to the response regulators of two-component regulatory systems and membrane translocators

“…In this model, there is a branch point of the signal relay, from which the regulatory cascade common to development and secondary metabolism is divided into two parts, one for the developmental process and the other for secondary metabolism. One of the proteins on the signal relay for streptomycin production after the branch point is an A-factor-responsive protein that binds to and probably activates the expression of strR, a regulatory gene in the streptomycin biosynthetic gene cluster (44). The amfgenes in the present study can be listed on the cascade for the developmental process downstream from the branch point.…”

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

“…Competitive EMSA, in which binding of AdpA to approximately 800 bp DNA fragments containing a 59 portion of each PAG with its upstream region was determined, showed that AdpA binds to 37 regions to activate 72 genes, including two known AdpA-dependent transcripts [strR-aphD (Vujaklija et al, 1991) and orf1-orf4 of AdBS2 (Yamazaki et al, 2004)]. The~800 bp DNA fragments were designed in such a way that they included a 600 bp sequence from 2500 to +100 (taking the first nucleotide of the start codon as +1).…”

Hormonal control by A-factor of morphological development and secondary metabolism in Streptomyces

“…6). 40),41) Concerning A-factor as a trigger of streptomycin biosynthesis, a question we raised was by what protein the A-factor signal was received and transmitted to the streptomycin biosynthesis genes. Analyses conducted by D. Vujaklija et al 42) on transcription of part of the streptomycin biosynthesis genes by S1 nuclease mapping showed that transcription of one mRNA species starting from the promoter of strR was …”