In Streptomyces coelicolor, bldA mutants are defective in antibiotic production and the development of aerial hyphae and spores. Subcloning analysis showed that sequences spanning an NcoI site in cloned bldA ÷ DNA were needed to allow complementation of a bldA mutant. Nucleotide sequencing revealed a tRNA-like sequence 9 bp downstream from the NcoI site. Five independent bldA mutations all fell in a 16-bp region in the tRNA-like sequence, one of them changing the putative anticodon. In RNA dot-blot analysis, hybridization was detected with a probe specific for the tRNA-like transcript but not with a probe for "anti-tRNA-like" transcripts. The transcripts detected were all in the sah-soluble RNA fraction and accumulated relatively late in growth. It is postulated that bldA specifies a tRNA that would recognize the codon UUA (for leucine). This codon is very rare in Streptomyces genes [which generally contain >70 mole% (G + C)], suggesting a possible role for bldA in translational control of development.

The anthelmintic drug levamisole causes hypercontraction of body wall muscles and lethality in nematode worms. In the nematode Caenorhabditis elegans, a genetic screen for levamisole resistance has identified 12 genes, three of which (unc-38, unc-29, and lev-1) encode nicotinic acetylcholine receptor (nAChR) subunits. Here we describe the molecular and functional characterization of another levamisole-resistant gene, unc-63, encoding a nAChR ␣ subunit with a predicted amino acid sequence most similar to that of UNC-38. Like UNC-38 and UNC-29, UNC-63 is expressed in body wall muscles. In addition, UNC-63 is expressed in vulval muscles and neurons. We also show that LEV-1 is expressed in body wall muscle, thus overlapping the cellular localization of UNC-63, UNC-38, and UNC-29 and suggesting possible association in vivo. This is supported by electrophysiological studies on body wall muscle, which demonstrate that a levamisole-sensitive nAChR present at the C. elegans neuromuscular junction requires both UNC-63 and LEV-1 subunits. Thus, at least four subunits, two ␣ types (UNC-38 and UNC-63) and two non-␣ types (UNC-29 and LEV-1), can contribute to levamisole-sensitive muscle nAChRs in nematodes.

The stringent response was elicited in the antibiotic producer Streptomyces coelicolor A3(2) either by amino acid depletion (nutritional shiftdown) or by the addition of serine hydroxamate; both led to increased levels of ppGpp and to a reduction in transcription from the four promoters of the rrnD rRNA gene set. Analysis of untreated batch cultures revealed elevated ppGpp levels at the end of exponential growth, preceding the onset of antibiotic production. The effect of provoking the stringent response on antibiotic production in exponentially growing cultures was assessed by S1 nuclease mapping of actIII, an early gene of the actinorhodin biosynthetic cluster. Expression of actIII occurred after nutritional shiftdown, but not after treatment with serine hydroxamate. Although the need for ppGpp in triggering antibiotic production remains equivocal, ppGpp synthesis alone does not appear to be sufficient to initiate secondary metabolism in S. coelicolor A3(2).

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