Metabolic initiation of differentiation and secondary metabolism by Streptomyces griseus: significance of the stringent response (ppGpp) and GTP content in relation to A factor

Abstract

I investigated the significance of the intracellular accumulation of guanosine 5'-diphosphate 3'-diphosphate (ppGpp) and of the coordinated decrease in the GTP pool for initiating morphological and physiological differentiation of Streptomyces griseus, a streptomycin-producing strain. In solid cultures, aerial mycelium formation was severely suppressed by the presence of excess nutrients. However, decoyinine, a specific inhibitor of GMP synthetase, enabled the cells to develop aerial mycelia in the suppressed cultures at concentrations which only partially inhibited growth. A factor (2S-isocapryloyl-3S-hydroxymethyl-,ybutyrolactone) added exogenously had no such effect. Decoyinine was also effective in initiating the formation of submerged spores in liquid culture. The ability to produce streptomycin did not increase but decreased drastically on the addition of decoyinine. This sharp decrease in streptomycin production was accompanied by a decrease in intracellular accumulation of ppGpp. A relaxed (rel) mutant was found among 25 thiopeptinresistant isolates which developed spontaneously. The rel mutant had a severely reduced ability to accumulate ppGpp during a nutritional shift-down and also during postexponential growth and showed a less extensive decrease in the GTP pool than that in the rel+ parental strain. The rel mutant failed to induce the enzymes amidinotransferase and streptomycin kinase, which are essential for the biosynthesis of streptomycin. The abilities to form aerial mycelia and submerged spores were still retained, but the amounts were less, and for both the onset of development was markedly delayed. The decreased ability to produce submerged spores was largely restored by the addition of decoyinine. This was accompained by an extensive GTP pool decrease. The rel mutant produced A factor normally, indicating that synthesis of A factor is controlled neither by ppGpp nor by GTP. Conversely, a mutant defective in A-factor synthesis accumulated as much ppGpp as did the parental strain. It was concluded that morphological differentiation of S. griseus results from a decrease in the pool of GTP, whereas physiological differentiation results from a more direct function of the rel gene product (ppGpp). It is also suggested that A factor may render the cell sensitive to receive and respond to the specified signal molecules, presumably ppGpp (for physiological differentiation) or GTP (for morphological differentiation).In bacteria, differentiation and induction of secondary metabolism start concomitantly in response to nutrient limitation. The study of differentiation in Streptomyces spp. has been especially of interest because this genus produces numerous antibiotics by secondary metabolism. The coupling of antibiotic production to morphological differentiation suggests a mechanistic connection which has not been demonstrated. The two processes do, however, often occur in parallel, (5). Typically, Streptomyces spp. develop aerial mycelia from substrate mycelia prior to the formation of spores (aerial sp...

Rare earth elements activate the secondary metabolite–biosynthetic gene clusters in Streptomyces coelicolor A3(2)

“…GYM medium was described previously. 16 Scandium chloride hexahydrate (ScCl 3 Á6H 2 O; purity, 99.9%) and lanthanum chloride heptahydrate (LaCl 3 Á7H 2 O; purity, 99.9%), together with other rare earths (all chloride-salts; purity, 497%) were purchased from Wako Pure Chemical (Osaka, Japan). These rare earths were added to the autoclaved agar medium after cooling to 50-60 1C.…”

Genus Kitasatospora, taxonomic features and diversity of secondary metabolites

“…Some Streptomyces strains produce SS but the SS are produced by fragmentation of mycelia (Figure 5b). [17][18][19][20][21][22][23] Feeding and shift-down experiments were carried out to identify the physiological regulation of SS formation of the strain KM-6054 T using minimum medium (MM) and casamino acid. 24 In feeding experiments, the strain produced SS when casamino acids were added in MM within 4 h after inoculation, but not when added after 6 h. In a nutritional shift-down experiment, SS were formed when casamino acids were removed from MM-CA after 8 h cultivation, but not when removed within 6 h. SS formation may therefore require at least 8 h incubation under nutrient-rich conditions.…”

“…Although reports about spores formed in the submerged culture are few, cases of Streptomyces griseus, 16,[21][22][23][24][25] Streptomyces roseosporus, 20 Streptomyces acrimycini, 18 Streptomyces albus 18 and Streptomyces venezuelae 19 have been published. Among them, Ochi et al 22,23 reported that the strain of S. griseus IFO 13189 forms SS along with a stringent response to nutritional starvation.…”

Activation of secondary metabolite–biosynthetic gene clusters by generating rsmG mutations in Streptomyces griseus

“…Spontaneous streptomycin-resistant mutants were obtained as colonies that grew within 5-7 days after spores were spread on glucose-yeast extract-malt extract (GYM) agar 11 containing various concentrations of streptomycin. Mutations in rpsL or rsmG genes were determined by DNA sequencing using the primers listed in Table 1.…”

“…Mutations in rpsL or rsmG genes were determined by DNA sequencing using the primers listed in Table 1. Cultivation was performed at 30 1C (for preculture) or 25 1C (for main culture) using SPY medium 11 with rotary shaking at 220 r.p.m.…”