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The uaY gene encodes the transcriptional activator of purine catabolism genes in Aspergillus nidulans. uaY12 results in strongly defective growth on purines as nitrogen sources and in strongly diminished transcription of UaY-regulated genes. This mutation introduces an ATG codon 64 bp upstream of the uaY ATG, generating a 68-codon open reading frame (uORFA), overlapping with the uaY ORF. uaY12 revertants fall into three categories: The majority eliminate the aberrant ATG. The growth and transcriptional phenotypes of these revertants are identical to those of the wild type. Two revertants create a stop codon in frame with the uaY12 aberrant ATG, shortening the length of the uORFA, thus uORFA no longer overlaps the uaY ORF. The latter are partial suppressors of the uaY12 mutation, while chain termination suppressors, in turn, suppress this novel phenotype. Two partial suppressors are unlinked to uaY. These two mutations result in a pleiotropic phenotype usually associated with ribosomal proteins. We hypothesize that uORFA strongly diminishes translation of the uaY ORF and that revertants negate this effect by a number of different mechanisms. The first-AUG rule and the phenomena of translational inhibition and leaky scanning provide a coherent explanation of the results presented in this article.

Spiramycin, a 16-membered macrolide antibiotic used in human medicine, is produced by Streptomyces ambofaciens; it comprises a polyketide lactone, platenolide, to which three deoxyhexose sugars are attached. In order to characterize the gene cluster governing the biosynthesis of spiramycin, several overlapping cosmids were isolated from an S. ambofaciens gene library, by hybridization with various probes (spiramycin resistance or biosynthetic genes, tylosin biosynthetic genes), and the sequences of their inserts were determined. Sequence analysis showed that the spiramycin biosynthetic gene cluster spanned a region of over 85 kb of contiguous DNA. In addition to the five previously described genes that encode the type I polyketide synthase involved in platenolide biosynthesis, 45 other genes have been identified. It was possible to propose a function for most of the inferred proteins in spiramycin biosynthesis, in its regulation, in resistance to the produced antibiotic or in the provision of extender units for the polyketide synthase. Two of these genes, predicted to be involved in deoxysugar biosynthesis, were inactivated by gene replacement, and the resulting mutants were unable to produce spiramycin, thus confirming their involvement in spiramycin biosynthesis. This work reveals the main features of spiramycin biosynthesis and constitutes a first step towards a detailed molecular analysis of the production of this medically important antibiotic.

Roundup® is a glyphosate-based herbicide (GBH) used worldwide both in agriculture and private gardens. Thus, it constitutes a substantial source of environmental contaminations, especially for water and soil, and may impact a number of non-target organisms essential for ecosystem balance. The soil filamentous fungus Aspergillus nidulans has been shown to be highly affected by a commercial formulation of Roundup® (R450), containing 450 g/L of glyphosate (GLY), at doses far below recommended agricultural application rate. In the present study, we used two-dimensional gel electrophoresis combined to mass spectrometry to analyze proteomic pattern changes in A. nidulans exposed to R450 at a dose corresponding to the no-observed-adverse-effect level (NOAEL) for macroscopic parameters (31.5 mg/L GLY among adjuvants). Comparative analysis revealed a total of 82 differentially expressed proteins between control and R450-treated samples, and 85% of them (70) were unambiguously identified. Their molecular functions were mainly assigned to cell detoxification and stress response (16%), protein synthesis (14%), amino acid metabolism (13%), glycolysis/gluconeogenesis/glycerol metabolism/pentose phosphate pathway (13%) and Krebs TCA cycle/acetyl-CoA synthesis/ATP metabolism (10%). These results bring new insights into the understanding of the toxicity induced by higher doses of this herbicide in the soil model organism A. nidulans. To our knowledge, this study represents the first evidence of protein expression modulation and, thus, possible metabolic disturbance, in response to an herbicide treatment at a dose that does not cause any visible effect. These data are likely to challenge the concept of 'substantial equivalence' when applied to herbicide-tolerant plants. [ABSTRACT FROM AUTHOR]

No gene coding for an adenine deaminase has been described in eukaryotes. However, physiological and genetical evidence indicates that adenine deaminases are present in the ascomycetes. We have cloned and characterised the genes coding for the adenine deaminases of Aspergillus nidulans, Saccharomyces cerevisiae and Schizosaccharomyces pombe. The A.nidulans gene was expressed in Escherichia coli and the purified enzyme shows adenine but not adenosine deaminase activity. The open reading frames coded by the three genes are very similar and obviously related to the bacterial and eukaryotic adenosine deaminases rather than to the bacterial adenine deaminases. The latter are related to allantoinases, ureases and dihydroorotases. The fungal adenine deaminases and the homologous adenosine deaminases differ in a number of residues, some of these being clearly involved in substrate specificity. Other prokaryotic enzymes in the database, while clearly related to the above, do not fit into either sub-class, and may even have a different specificity. These results imply that adenine deaminases have appeared twice in the course of evolution, from different ancestral enzymes constructed both around the alpha/beta barrel scaffold.