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L Riles

Publications and source records attributed to L Riles.

13 recordsLinked to original sources
PubMed2000

A chemical genomics approach toward understanding the global functions of the target of rapamycin protein (TOR).

The target of rapamycin protein (TOR) is a highly conserved ataxia telangiectasia-related protein kinase essential for cell growth. Emerging evidence indicates that TOR signaling is highly complex and is involved in a variety of cellular processes. To understand its general functions, we took a chemical genomics approach to explore the genetic interaction between TOR and other yeast genes on a genomic scale. In this study, the rapamycin sensitivity of individual deletion mutants generated by the Saccharomyces Genome Deletion Project was systematically measured. Our results provide a global view of the rapamycin-sensitive functions of TOR. In contrast to conventional genetic analysis, this approach offers a simple and thorough analysis of genetic interaction on a genomic scale and measures genetic interaction at different possible levels. It can be used to study the functions of other drug targets and to identify novel protein components of a conserved core biological process such as DNA damage checkpoint/repair that is interfered with by a cell-permeable chemical compound.

Cell Cycle Proteins↗
PubMed1999

Functional characterization of the S. cerevisiae genome by gene deletion and parallel analysis.

The functions of many open reading frames (ORFs) identified in genome-sequencing projects are unknown. New, whole-genome approaches are required to systematically determine their function. A total of 6925 Saccharomyces cerevisiae strains were constructed, by a high-throughput strategy, each with a precise deletion of one of 2026 ORFs (more than one-third of the ORFs in the genome). Of the deleted ORFs, 17 percent were essential for viability in rich medium. The phenotypes of more than 500 deletion strains were assayed in parallel. Of the deletion strains, 40 percent showed quantitative growth defects in either rich or minimal medium.

Culture Media↗
PubMed1999

Systematic analysis of S. cerevisiae chromosome VIII genes.

To begin genome-wide functional analysis, we analysed the consequences of deleting each of the 265 genes of chromosome VIII of Saccharomyces cerevisiae. For 33% of the deletion strains a growth phenotype could be detected: 18% of the genes are essential for growth on complete glucose medium, and 15% grow significantly more slowly than the wild-type strain or exhibit a conditional phenotype when incubated under one of 20 different growth conditions. Two-thirds of the mutants that exhibit conditional phenotypes are pleiotropic; about one-third of the mutants exhibit only one phenotype. We also measured the level of expression directed by the promoter of each gene. About half of the promoters direct detectable transcription in rich glucose medium, and most of these exhibited only low or medium activity. Only 1% of the genes are expressed at about the same level as ACT1. The number of active promoters increased to 76% upon growth on a non-fermentable carbon source, and to 93% in minimal glucose medium. The majority of promoters fluctuated in strength, depending on the medium.

DNA Primers↗
PubMed1997

Genetic and physical maps of Saccharomyces cerevisiae.

Genetic and physical maps for the 16 chromosomes of Saccharomyces cerevisiae are presented. The genetic map is the result of 40 years of genetic analysis. The physical map was produced from the results of an international systematic sequencing effort. The data for the maps are accessible electronically from the Saccharomyces Genome Database (SGD: http://genome-www.stanford. edu/Saccharomyces/).

Chromosome Mapping↗
PubMed1997

The nucleotide sequence of Saccharomyces cerevisiae chromosome XII.

The yeast Saccharomyces cerevisiae is the pre-eminent organism for the study of basic functions of eukaryotic cells. All of the genes of this simple eukaryotic cell have recently been revealed by an international collaborative effort to determine the complete DNA sequence of its nuclear genome. Here we describe some of the features of chromosome XII.

Base Sequence↗
PubMed1997

The sequence of a nearly unclonable 22.8 kb segment on the left arm chromosome VII from Saccharomyces cerevisiae reveals ARO2, RPL9A, TIP1, MRF1 genes and six new open reading frames.

The nucleotide sequence of 22,803 bp on the left arm of chromosome VII was determined by polymerase chain reaction-based approaches to compensate for the unstable character of cosmid clones from this region of the chromosome. The coding density of the sequence is particularly high (more than 83%). Twelve open reading frames (ORFs) longer than 300 bp were found, two of which (at the left side) have been described previously (James et al., 1995) after sequencing of an overlapping cosmid. Four other ORFs correspond to published sequences of the known genes ARO2, RPL9A, TIP1 and MRF1. ARO2 codes for chorismate synthetase. RPL9A for protein L9 of the large ribosomal subunit and MRF1 for a mitochondrial translation release factor. The TIP1 product interacts with Sec20p and is thus involved in transport from endoplasmic reticulum to Golgi. Five of the remaining ORFs have not been identified previously, while the sixth (YGL142c) has been partially sequenced as it lies 5' upstream of MRF1. These six ORFs are relatively large (between 933 and 3657 nucleotides). YGL146c, YGL142c, YGL140c and YGL139w have no significant homology to any protein sequence presently available in the public databases, but show two, nine, nine and eight putative transmembrane spans, respectively. YGL144c has a serine active site signature of lipases. YGL141w has limited homology to several human proteins, one of which mediates complex formation between papillomavirus E6 oncoprotein and tumor suppressor protein p53.

Arabidopsis Proteins↗
PubMed1996

Green fluorescent protein as a marker for gene expression and subcellular localization in budding yeast.

The green fluorescent protein (GFP) from the jellyfish Aequorea victoria has attracted much attention as a tool to study a number of biological processes. This study describes the use of GFP as a vital reporter molecule for localization and expression studies in Saccharomyces cerevisiae. Construction of GFP expression vectors which allow N- or C-terminal fusion of the gfp gene to a gene of interest allowed the generation of fusion proteins whose subcellular localization was followed by fluorescence microscopy in living yeast cells. Analysis of three unknown open reading frames obtained from the budding yeast chromosome XIV resulted in distinct staining patterns, allowing prediction of the cellular localization of these unknown proteins. Furthermore, GFP was used to construct a gene replacement cassette which, after homologous integration into the genomic locus, placed the gfp gene behind a promoter of interest. The amount of GFP produced from this promoter was then quantified in living yeast cells by flow cytometry. With this novel replacement cassette a gene of interest can be deleted and at the same time its expression level studied under various growth conditions. The experiments presented here suggest that GFP represents a convenient fluorescent marker for localization studies as well as gene expression studies in budding yeast. Systematic studies of a large number of genes should benefit from such assays.

Animals↗
PubMed1994

Mapping of DBR1 and YPK1 suggests a major revision of the genetic map of the left arm of Saccharomyces cerevisiae Chromosome XI.

The Saccharomyces cerevisiae dbr1 mutation has been mapped on the left arm of chromosome XI. XIL is a chromosome arm that was until now rather sparsely populated with accurately mapped markers. On the basis of physical data, the overall order of markers is inverted relative to the existing genetic map of XI. We present tetrad analyses using a variety of markers on XI that indicate that the existing genetic map of XIL should be inverted, at least for the strains in which our mapping was carried out, and probably for other S. cerevisiae strains.

Chromosome Mapping↗
PubMed1993

Physical maps of the six smallest chromosomes of Saccharomyces cerevisiae at a resolution of 2.6 kilobase pairs.

Physical maps of the six smallest chromosomes of Saccharomyces cerevisiae are presented. In order of increasing size, they are chromosomes I, VI, III, IX, V and VIII, comprising 2.49 megabase pairs of DNA. The maps are based on the analysis of an overlapping set of lambda and cosmid clones. Overlaps between adjacent clones were recognized by shared restriction fragments produced by the combined action of EcoRI and HindIII. The average spacing between mapped cleavage sites is 2.6 kb. Five of the six chromosomes were mapped from end to end without discontinuities; a single internal gap remains in the map of chromosome IX. The reported maps span an estimated 97% of the DNA on the six chromosomes; nearly all the missing segments are telomeric. The maps are fully cross-correlated with the previously published SfiI/NotI map of the yeast genome by A. J. Link and M. V. Olson. They have also been cross-correlated with the yeast genetic map at 51 loci.

Base Sequence↗
PubMed1988

Nonsense mutations in essential genes of Saccharomyces cerevisiae.

A new method for isolating nonsense mutations in essential yeast genes has been used to develop a collection of 115 ochre mutations that define 94 complementation groups. The mutants are isolated in a genetic background that includes an ochre suppressor on a metastable plasmid and a suppressible colony-color marker on a chromosome. When the parental strain is plated on a rich medium, the colonies display a pattern of red, plasmid-free sectors on a white background. Mutants containing an ochre mutation in any essential yeast gene give rise to nonsectoring, white colonies, since cell growth is dependent on the presence of the plasmid-borne suppressor. Analysis of the data suggests that mutations are being recovered from a pool of approximately 250 genes.

Genes, Fungal↗