New sporulation loci in Streptomyces coelicolor A3(2)

Sporulation mutants of Streptomyces coelicolor appear white because they ar e defective in the synthesis of the grey polyketide spore pigment, and such white (whi) mutants had been used to define eight sporulation loci, whiA, whiB, whiD, whiE, whiG, whiH, whiI, and whiJ (K. F. Chater, J. Gen. Microbi ol. 72:9-28, 1972; N. J. Ryding, Ph.D. thesis, University of East Anglia, 1 995). In an attempt to identify new whi loci, we mutagenized S. coelicolor M145 spores with nitrosoguanidine and identified 770 mutants with colonies ranging from white to medium grey. After excluding unstable strains, we exa mined the isolates by phase-contrast microscopy and chose 115 whi mutants w ith clear morphological phenotypes for further study. To exclude mutants re presenting cloned whi genes, self-transmissible SCP2*-derived plasmids carr ying whiA, whiB, whiG, whiH, or whiJ (but not whiD, whiE, or whiI) were int roduced into each mutant by conjugation, and strains in which the wild-type phenotype was restored either partially or completely by any of these plas mids were excluded from further analysis. In an attempt to complement some of the remaining 31 whi mutants, an SCP2* library of wild-type S. coelicolo r chromosomal DNA was introduced into 19 of the mutants by conjugation. Clo nes restoring the wild-type phenotype to 12 of the 19 strains were isolated and found to represent five distinct loci, designated whiK, whiL, whiM, wh iN, and whiO. Each of the five loci was located on the ordered cosmid libra ry: whit, whiM, whiN, and whiO occupied positions distinct from previously cloned whi genes; whiK was located on the same cosmid overlap as whiD, but the two loci were shown by complementation to be distinct. The phenotypes r esulting from mutations at each of these new loci are described.