A Purine-Related Metabolite Negatively Regulates fixNOQP Expression in Sinorhizobium meliloti by Modulation of fixK Expression

5-aminoimidazole-4-carboxamide nucleotide (AICAR) is a negative effector of cytochrome terminal oxidase cbb3 production in Rhizobium etli. In this work, the effect of AICAriboside (AICAr), the precursor of AICAR on the expression of the Sinorhizobium meliloti fixNOQP operon encoding the symbiotic terminal oxidase cbb3, was analyzed. AICAr reduced the microaerobic induction levels of fixN-lacZ and fixT-lacZ gene fusions 18- and sevenfold respectively, and both genes were activated by the transcriptional activator FixK. A fixK-lacZ fusion presented 14-fold-reduced induction levels in microaerobic cell cultures in the presence of AICAr. AICAr also reduced three-fold the microaerobic expression levels of the nifA-lacZ fusion, whose expression as well as that of fixK is controlled by the two-component system FixL-FixJ. In contrast, AICAr had no effect on the expression levels of a hemA-lacZ fusion. These data suggest that AICAr prevents fixNOQP induction by the inhibition of fixK transcription.

Extragenic suppressors of Saccharomyces cerevisiae prp4 mutations identify a negative regulator of PRP genes.

The PRP4 gene encodes a protein that is a component of the U4/U6 small nuclear ribonucleoprotein particle and is necessary for both spliceosome assembly and pre-mRNA splicing. To identify genes whose products interact with the PRP4 gene or gene product, we isolated second-site suppressors of temperature-sensitive prp4 mutations. We limited ourselves to suppressors with a distinct phenotype, cold sensitivity, to facilitate analysis of mutants. Ten independent recessive suppressors were obtained that identified four complementation groups, spp41, spp42, spp43 and spp44 (suppressor of prp4, numbers 1-4). spp41-spp44 suppress the pre-mRNA splicing defect as well as the temperature-sensitive phenotype of prp4 strains. Each of these spp mutations also suppresses prp3; spp41 and spp42 suppress prp11 as well. Neither spp41 nor spp42 suppressors null alleles of prp3 or prp4, indicating that the suppression does not occur via a bypass mechanism. The spp41 and spp42 mutations are neither allele- nor gene-specific in their pattern of suppression and do not result in a defect in pre-mRNA splicing. Thus the SPP41 and SPP42 gene products are unlikely to participate directly in mRNA splicing or interact directly with Prp3p or Prp4p. Expression of PRP3-lacZ and PRP4-lacZ gene fusions is increased in spp41 strains, suggesting that wild-type Spp41p represses expression of PRP3 and PRP4. SPP41 was cloned and sequenced and found to be essential. spp43 is allelic to the previously identified suppressor srn1, which encodes a negative regulator of gene expression.