WhiD and WhiB, homologous proteins required for different stages of sporulation in Streptomyces coelicolor A3(2)

The whiD locus, which is required for the differentiation of Streptomyces c oelicolor aerial hyphae into mature spore chains, was localized by map-base d cloning to the overlap between cosmids 6G4 and D63 of the minimal ordered library of Redenbach et al, (M, Redenbach et al., Mol. Microbiol. 21:77-96 , 1996), Subcloning and sequencing showed that whiD encodes a homologue of WhiB, a protein required for the initiation of sporulation septation in S. coelicolor. WhiD and WhiB belong to a growing family of small (76- to 112-r esidue) proteins of unknown biochemical function in which four cysteines ar e absolutely conserved; all known members of this family are found in the a ctinomycetes. A constructed whiD null mutant showed reduced levels of sporu lation, and those spores that did farm were heat sensitive, lysed extensive ly, and were highly irregular in size, arising at least in part from irregu larity in septum placement. The whiD null mutant showed extreme variation i n spore cell wall deposition; most spores had uniformly thin (20- to 30-nm) walls, but spore chains were frequently observed in which there was irregu lar but very pronounced (up to 170 nm) fell wall thickening at the junction s between spores. whiD null mutant spores were frequently partitioned into irregular smaller units through the deposition of additional septa, which w ere often laid down in several different planes, very close to the spore po les. These "minicompartments" appeared to be devoid of chromosomal DNA. Two whiD promoters, whiDp1 and whiDp2, were identified, and their activities w ere analyzed during development of wild-type S. coelicolor on solid medium, Both promoters were developmentally regulated; whiDp1 and whiDp2 transcrip ts were detected transiently, approximately at the time when sporulation se pta were observed in the aerial hyphae.