# The Link between Purine Metabolism and Production of Antibiotics in _Streptomyces_
## 2 Stringent Response and Morphological Differentiation
Two proteins, RelA and RSH, participate in (p)ppGpp metabolism in _Streptomyces_ spp., likely with a division of labor in which RelA and RSH respond to amino acid/glucose starvation and phosphate starvation, respectively [20; 21; 22]. Starvation of _Streptomyces_ spp. results in elevated levels of (p)ppGppas well as a reduced GTP pool, as also seen in _B. subtilis_, accompanied by reduced transcription of rRNA genes [23; 24; 25; 26]. For example, _S. clavuligerus_ in which _relA_ is inactivated fails to synthesize (p)ppGpp and maintains steady levels of GTP during growth, whereas wild-type cells exhibit an approximately three-fold decrease in GTP levels as (p)ppGpp accumulates [27]. A similar (p)ppGpp-dependent decrease in cellular GTP levels was previously demonstrated in _S. griseus_ during nutritional shift-down, and it was attributed to ppGpp-mediated inhibition of IMP dehydrogenase (GuaB; Figure 1) [28]. Starvation also elicits a complex morphological differentiation in Streptomyces, a process that involves formation of aerial hyphae and unicellular spores for dispersal, and this differentiation may be accompanied by production of bioactive secondary metabolites. Erection of aerial hyphae requires the _bld_ (bald) gene products, while differentiation of aerial hyphae into spores requires the _vni_ (white) gene products [29]. The transcriptional regulator BldD controls a large number of genes, many of which are involved in differentiation and antibiotic production [3]. The secondary messenger c-di-GMP, which is synthesized from two molecules of GTP by diguanylate cyclases and degraded by phosphodiesterase, also participates in differentiation. The two _S. coelicolor_ proteins RmdA and RmdB possess phosphodiesterase activity and the _rmdA-rmdB_ double mutant accumulates three-fold higher levels of c-di-GMP and is completely deficient in aerial hyphae formation [30]. Deficiency in generation of aerial mycelium is also a characteristic of _S. coelicolor_ deleted for _cdgB_, which encodes a diguanylate cyclase, and of cells over-expressing _cdgB_; _cdgB_ over-expression also inhibits formation of the antibiotic actinorhodin [31]. Expression of _cdgB_ is controlled by BldD; notably, DNA binding by BldD is activated by direct binding of c-di-GMP, leading to repression of the BldD-controlled sporulation genes during vegetative growth [32]. Figure 1: Overview of purine metabolism. The _de novo_ biosynthesis (turquoise) starts with phosphoribosyl pyrophosphate (PRPP) and ends with inosine monophosphate (IMP); the intermediate AICAR (5-aminoimidazole-4-carboxamide ribonucleotide) accumulates in presence of dihydrofolate reductase inhibitors [33]. Intermediates in salvage pathways are outlined in purple. In some species, guanine, xanthine, and hypoxanthine are converted to the corresponding mononucleotides by different enzymes. Xdh can initiate degradation of purines (gray). RelA/SpoT Homolog (RSH) family enzymes are responsible for (p)ppGpp synthesis and hydrolysis, while diguanylate cyclase (DGC) synthesizes c-di-GMP (red).