By using a PCR approach based on conserved regions of ADP-glucose pyrophosphorylases, a glgC gene was cloned from Streptomyces coelicolor A3(2). The deduced glgC gene product showed end-to-end relatedness to other bacterial ADP-glucose pyrophosphorylases. The glgC gene is about 1,000 kb from the leftmost chromosome end and is not closely linked to either of the two glgB genes of S. coelicolor, which encode glycogen branching enzymes active in different locations in differentiated colonies. Disruption of glgC eliminated only the first of two temporal peaks of ADP-glucose pyrophosphorylase activity and glycogen accumulation and prevented cytologically observable glycogen accumulation in the substrate mycelium of colonies (phase I), while glycogen deposition in young spore chains (phase II) remained readily detectable. The cloned glgC gene therefore encodes an ADP-glucose pyrophosphorylase essential only for phase I (and it is therefore named glgCI). A second, phase II-specific, glgC gene should also exist in S. coelicolor, though it was not detected by hybridization analysis.Glycogen is a branched homopolysaccharide of ␣-1,4-linked glucose subunits with ␣-1,6-linked glucose at the branching points. Many bacteria accumulate glycogen when their growth is nutrient limited in the presence of an excess of carbon source (40). The biosynthesis of bacterial glycogen from glucose-1-phosphate is catalyzed by three enzymes: ADP-glucose pyrophosphorylase, glycogen synthase, and branching enzyme, the products of the glgC, glgA, and glgB genes, respectively (38, 40). These genes, together with glgY (ϭ glgP), coding for glycogen phosphorylase, and glgX, which probably encodes a glycogen debranching enzyme, are organized in two adjacent operons, glgBX and glgCAY in Escherichia coli (38). In E. coli, glycogen synthesis is regulated at the enzymatic level by allosteric regulation of ADP-glucose pyrophosphorylase and at the mRNA abundance level by positively acting factors, such as cyclic AMP, ppGpp, and the stationary-phase factor s (see references 28, 38, and 41 for recent reviews), and by the negatively acting product of a pleiotropic carbon storage regulatory gene, csrA (27,50). In the developmentally more complex organism Bacillus subtilis, intriguingly, a single glg operon containing all the essential glg genes is apparently controlled by a sporulation mother-cell-specific form of RNA polymerase (E E ), suggesting that glycogen metabolism is developmentally and spatially controlled in B. subtilis (22).Streptomyces spp. are gram-positive bacteria that undergo particularly complex morphological differentiation. Initially, the spores germinate to form a highly branched substrate mycelium, which is firmly attached to the solid medium. Later, the aerial mycelium emerges, and spores are generated by septation of the aerial hyphae (14,15, 31,32,49). During Streptomyces differentiation, glycogen accumulates in two temporally and spatially distinct phases (4, 6, 37). The first phase (phase I) takes place in substrate hyphae around the ...