Genetic Analysis of A-factor Synthesis in Streptomyces coelicolor A3(2) and Streptomyces griseus

Abstract

A-factor is a potent pleiotropic effector produced by Streptomyces griseus and is essential for streptomycin production and spore formation in this organism. Its production is widely distributed among various actinomycetes including Streptomyces coelicolor A3(2). Genetic analysis of A-factor production was carried out with S . coelicolor A3(2), and two closely linked loci for A-factor mutations (afsA and B) were identified between cysD and leuB on the chromosomal linkage map. In contrast, genetic crosses of A-factor-negative mutants of S. griseus, using a protoplast fusion technique, failed to give a fixed locus for A-factor gene(s) and suggested involvement of an extrachromosomal or transposable genetic element in A-factor synthesis in this organism. INTRODUCTIONA-factor, 2-S-isocapryloyl-3-S-hydroxymethyl-y-butyrolactone, found in Streptomyces griseus and Streptomyces bikiniensis (Khokhlov et al., 1973), is a potent autoregulating factor essential for both streptomycin biosynthesis and spore formation in these organisms (Khokhlov et al., 1973;Khokhlov, 1980; Hara & Beppu, 1982a). Furthermore, recent work from this laboratory has revealed that A-factor is also involved in streptomycin resistance in these organisms through its inducible effect on the synthesis of streptomycin-6-phosphotransferase (Hara & Beppu, 1982 b). Mutants lacking A-factor simultaneously lose streptomycin production capability, the ability to form spores and a large degree of streptomycin resistance. Exogenous supplementation of A-factor to the culture of these mutants restores all these phenotypes. A-factor-negative mutants of S . griseus and S. bikiniensis were easily obtained at high frequency by treatment with acridine dyes or by incubation at high temperature, which suggested involvement of an extrachromosomal genetic determinant in the biosynthesis of A-factor in these organisms.A-factor productivity has been found in various species of actinomycetes including Streptomyces coelicolor A3(2) (Efremenkova et al., 1979; Hara & Beppu, 1982a), but in contrast to S . griseus and S. bikiniensis, no A-factor-negative mutants of S. coelicolor A3(2) were obtained by the curing treatments. In this report, we describe genetic analyses of A-factor genes of S . coelicolor A3(2) and S . griseus, using conjugation and protoplast fusion techniques, respectively. The results clearly indicate that A-factor gene(s) of S. coelicofor A3(2) are located at a fixed position on the chromosomal linkage map, while in S . griseus, they appear to be extrachromosomal.t Present address: Fermentation Technology Laboratories, Meiji

A combined genetic and physical map of the Streptomyces coelicolor A3(2) chromosome

“…(iv) A cloned fragment isolated on the basis of its hybridization to dnaA of Pseudomonas putida and therefore likely to lie near a chromosomal origin of replication (30) was mapped to a position (on AseI fragment H) at about 8 o'clock; the conclusion that this DNA includes an origin of replication of the S. coelicolor chromosome is established by the recent work of Calcutt and Schmidt (14) and Zakrzewska-Czerwinska and Schrempf (92). (v) The afsR gene (46) was originally cloned by its ability to stimulate antibiotic production in S. lividans and was then found to complement an afsB mutation, which had been mapped genetically close to cysD (33). However, Stein and Cohen (84) showed by a genetic argument that the cloned DNA did not represent the wild-type allele of afsB.…”

The regulation of antibiotic production in Streptomyces coelicolor A3(2)

“…as indicators of cell density akin to the homoserine lactone derivatives made by Vibrio species (Fuqua e t al., 1994;Williams, 1994). Similar observations were made for the accumulation of virginae butanolide C that stimulates virginamycin production in Streptomyes virginiae (Y ang e t al., 1995), and for A-factor production by S. grisezls (Hara & Beppu, 1982). It thus appears more likely that the ybutyrolactones are produced in response to certain physiological or environmental signals.…”

A relA/spoT Homologous Gene from Streptomyces coelicolor A3(2) Controls Antibiotic Biosynthetic Genes

“…Neither actinorhodine nor undecylprodigiosin (as well as reduced amounts of methylenomycin and CDA) could be detected in afsB mutants (17), which were suppressed by the afsR gene (18,19). In abaA-ORFB (20) mutants, actinorhodine and undecylprodigiosin production is blocked, while the level of CDA is reduced and methylenomycin remains unaffected.…”