Biomedical subjects

Linda Riles

Publications and source records attributed to Linda Riles.

4 recordsLinked to original sources

PubMed2005

Discovery, validation, and genetic dissection of transcription factor binding sites by comparative and functional genomics.

Completing the annotation of a genome sequence requires identifying the regulatory sequences that control gene expression. To identify these sequences, we developed an algorithm that searches for short, conserved sequence motifs in the genomes of related species. The method is effective in finding motifs de novo and for refining known regulatory motifs in Saccharomyces cerevisiae. We tested one novel motif prediction of the algorithm and found it to be the binding site of Stp2; it is significantly different from the previously predicted Stp2 binding site. We show that Stp2 physically interacts with this sequence motif, and that stp2 mutations affect the expression of genes associated with the motif. We demonstrate that the Stp2 binding site also interacts genetically with Stp1, a regulator of amino acid permease genes and, with Sfp1, a key regulator of cell growth. These results illuminate an important transcriptional circuit that regulates cell growth through external nutrient uptake.

Arylsulfotransferase↗

PubMed2004

Large-scale screening of yeast mutants for sensitivity to the IMP dehydrogenase inhibitor 6-azauracil.

Mutations in several genes encoding components of the RNA polymerase II elongation machinery render S. cerevisiae cells sensitive to the drug 6-azauracil (6AU), an inhibitor of IMP dehydrogenase and orotidylate decarboxylase. It is thought that a reduction in nucleotide levels following drug treatment causes transcriptional elongation to be more dependent on a fully functional RNA polymerase. To gain insight into the basis of the 6AU-sensitive phenotype and discern its specificity, we screened almost 3000 deletion mutants for growth in the presence of drug; 42 (1.5%) were reproducibly sensitive to the drug. The sensitive mutants included several missing known transcription elongation factors, but the majority were in genes involved in other cellular processes. Not all of the 6AU-sensitive strains displayed cross-sensitivity to mycophenolic acid (MPA), another drug that inhibits IMP dehydrogenase and has been employed as a screening agent for elongation mutants, showing that these two drugs are mechanistically distinct. Several of the mutants were tested for the ability to induce transcription of IMP dehydrogenase-encoding genes, in response to 6-AU and MPA treatment. As expected, mutants defective in transcriptional elongation factors were unable to fully induce IMPDH expression. However, most of the 6AU-sensitive strains had normal levels of IMPDH expression. Thus, although 6AU-sensitivity often results from defects in the elongation machinery, mutations that compromise processes other than transcription and induction of IMPDH also lead to sensitivity to this drug.

Blotting, Northern↗

PubMed2002

Functional profiling of the Saccharomyces cerevisiae genome.

Determining the effect of gene deletion is a fundamental approach to understanding gene function. Conventional genetic screens exhibit biases, and genes contributing to a phenotype are often missed. We systematically constructed a nearly complete collection of gene-deletion mutants (96% of annotated open reading frames, or ORFs) of the yeast Saccharomyces cerevisiae. DNA sequences dubbed 'molecular bar codes' uniquely identify each strain, enabling their growth to be analysed in parallel and the fitness contribution of each gene to be quantitatively assessed by hybridization to high-density oligonucleotide arrays. We show that previously known and new genes are necessary for optimal growth under six well-studied conditions: high salt, sorbitol, galactose, pH 8, minimal medium and nystatin treatment. Less than 7% of genes that exhibit a significant increase in messenger RNA expression are also required for optimal growth in four of the tested conditions. Our results validate the yeast gene-deletion collection as a valuable resource for functional genomics.

Cell Size↗

PubMed2002

Gene disruption.

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Gene Duplication↗