Members of the soil‐dwelling prokaryotic genus Streptomyces produce many secondary metabolites, including antibiotics and anti‐tumour agents. Their formation is coupled with the onset of development, which is triggered by the nutrient status of the habitat. We propose the first complete signalling cascade from nutrient sensing to development and antibiotic biosynthesis. We show that a high concentration of N‐acetylglucosamine—perhaps mimicking the accumulation of N‐acetylglucosamine after autolytic degradation of the vegetative mycelium—is a major checkpoint for the onset of secondary metabolism. The response is transmitted to antibiotic pathway‐specific activators through the pleiotropic transcriptional repressor DasR, the regulon of which also includes all N‐acetylglucosamine‐related catabolic genes. The results allowed us to devise a new strategy for activating pathways for secondary metabolite biosynthesis. Such ‘cryptic’ pathways are abundant in actinomycete genomes, thereby offering new prospects in the fight against multiple drug‐resistant pathogens and cancers.

Haydon and Guest (Haydon, D. J, and Guest, J. R. (1991) FEMS Microbiol. Lett. 63, 291-295) first described the helix-turn-helix GntR family of bacterial regulators. They presented them as transcription factors sharing a similar N-terminal DNA-binding (d-b) domain, but they observed near-maximal divergence in the C-terminal effector-binding and oligomerization (E-b/O) domain. To elucidate this C-terminal heterogeneity, structural, phylogenetic, and functional analyses were performed on a family that now comprises about 270 members. Our comparative study first focused on the C-terminal E-b/O domains and next on DNA-binding domains and palindromic operator sequences, has classified the GntR members into four subfamilies that we called FadR, HutC, MocR, and YtrA. Among these subfamilies a degree of similarity of about 55% was observed throughout the entire sequence. Structure/function associations were highlighted although they were not absolutely stringent. The consensus sequences deduced for the DNA-binding domain were slightly different for each subfamily, suggesting that fusion between the D-b and E-b/O domains have occurred separately, with each subfamily having its own D-b domain ancestor. Moreover, the compilation of the known or predicted palindromic cis-acting elements has highlighted different operator sequences according to our subfamily subdivision. The observed C-terminal E-b/O domain heterogeneity was therefore reflected on the DNA-binding domain and on the cis-acting elements, suggesting the existence of a tight link between the three regions involved in the regulating process.

Summary: Members of the soil‐dwelling, sporulating prokaryotic genus Streptomyces are indispensable for the recycling of the most abundant polysaccharides on earth (cellulose and chitin), and produce a wide range of antibiotics and industrial enzymes. How do these organisms sense the nutritional state of the environment, and what controls the signal for the switch to antibiotic production and morphological development? Here we show that high extracellular concentrations of N‐acetylglucosamine, the monomer of chitin, prevent Streptomyces coelicolor progressing beyond the vegetative state, and that this effect is absent in a mutant defective of N‐acetylglucosamine transport. We provide evidence that the signal is transmitted through the GntR‐family regulator DasR, which controls the N‐acetylglucosamine regulon, including the pts genes ptsH, ptsI and crr needed for uptake of N‐acetylglucosamine. Deletion of dasR or the pts genes resulted in a bald phenotype. Binding of DasR to its target genes is abolished by glucosamine 6‐phosphate, a central molecule in N‐acetylglucosamine metabolism. Extracellular complementation experiments with many bld mutants showed that the dasR mutant is arrested at an early stage of the developmental programme, and does not fit in the previously described bld signalling cascade. Thus, for the first time we are able to directly link carbon (and nitrogen) metabolism to development, highlighting a novel type of metabolic regulator, which senses the nutritional state of the habitat, maintaining vegetative growth until changing circumstances trigger the switch to sporulation. Our work, and the model it suggests, provide new leads towards understanding how microorganisms time developmental commitment.

N-acetylglucosamine, the monomer of chitin, is a favored carbon and nitrogen source for streptomycetes. Its intracellular catabolism requires the combined actions of the N-acetylglucosamine-6-phosphate (GlcNAc-6P) deacetylase NagA and the glucosamine-6-phosphate (GlcN-6P) deaminase/isomerase NagB. GlcNAc acts as a signaling molecule in the DasR-mediated nutrient sensing system, activating development and antibiotic production under poor growth conditions (famine) and blocking these processes under rich conditions (feast). In order to understand how a single nutrient can deliver opposite information according to the nutritional context, we carried out a mutational analysis of the nag metabolic genes nagA, nagB, and nagK. Here we show that the nag genes are part of the DasR regulon in Streptomyces coelicolor, which explains their transcriptional induction by GlcNAc. Most likely as the result of the intracellular accumulation of GlcN-6P, nagB deletion mutants fail to grow in the presence of GlcNAc. This toxicity can be alleviated by the additional deletion of nagA. We recently showed that in S. coelicolor, GlcNAc is internalized as GlcNAc-6P via the phosphoenolpyruvate-dependent sugar phosphotransferase system (PTS). Considering the relevance of GlcNAc for the control of antibiotic production, improved insight into GlcNAc metabolism in Streptomyces may provide new leads toward biotechnological applications.