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Brenda Andrews

Publications and source records attributed to Brenda Andrews.

27 records · Page 2Linked to original sources
PubMed2003

Late-G1 cyclin-CDK activity is essential for control of cell morphogenesis in budding yeast.

The accurate spatial and temporal coordination of cell polarization with DNA replication and segregation guarantees the fidelity of genetic transmission. Here we report that in Saccharomyces cerevisiae, a build-up or burst of G1 cyclin-dependent kinase (CDK) activity through activation of the cyclin genes CLN1,2 and PCL1,2 is essential for cell morphogenesis, but not for other events associated with the G1-S-phase transition, including DNA replication. Strains lacking a burst of late-G1 cyclin-CDK activity (LG1C(-)) undergo a catastrophic morphogenesis and halt the nuclear cycle at the morphogenesis checkpoint in G2 phase. Consistent with a role in morphogenesis, the Pho85 G1 cyclins Pcl1 and Pcl2 show a unique pattern of localization to sites of polarized cell growth, and strains lacking PCL1 and PCL2 show genetic interactions with the cell polarity GTPase Cdc42, its regulators and downstream effectors. Our data suggest that inability to assemble a septin ring and localize the GTP exchange factor Cdc24 at the incipient bud site may be the primary morphogenetic defects in LG1C-depleted cells. We conclude that a burst of late G1 cyclin-CDK activity is essential for establishing cell polarity and development of the cleavage apparatus.

Cell Cycle↗
PubMed2003

Regulation of the yeast amphiphysin homologue Rvs167p by phosphorylation.

The yeast amphiphysin homologue Rvs167p plays a role in regulation of the actin cytoskeleton, endocytosis, and sporulation. Rvs167p is a phosphoprotein in vegetatively growing cells and shows increased phosphorylation upon treatment with mating pheromone. Previous work has shown that Rvs167p can be phosphorylated in vitro by the cyclin-dependent kinase Pho85p complexed with its cyclin Pcl2p. Using chymotryptic phosphopeptide mapping, we have identified the sites on which Rvs167p is phosphorylated in vitro by Pcl2p-Pho85p. We have shown that these same sites are phosphorylated in vivo during vegetative growth and that phosphorylation at two of these sites is Pcl-Pho85p dependent. In cells treated with mating pheromone, the MAP kinase Fus3p is needed for full phosphorylation of Rvs167p. Functional genomics and genetics experiments revealed that mutation of other actin cytoskeleton genes compromises growth of a strain in which phosphorylation of Rvs167p is blocked by mutation. Phosphorylation of Rvs167p inhibits its interaction in vitro with Las17p, an activator of the Arp2/3 complex, as well as with a novel protein, Ymr192p. Our results suggest that phosphorylation of Rvs167p by a cyclin-dependent kinase and by a MAP kinase is an important mechanism for regulating protein complexes involved in actin cytoskeleton function.

Actin-Related Protein 2↗
PubMed2003

Functional genomics and proteomics: charting a multidimensional map of the yeast cell.

The challenge of large-scale functional genomics projects is to build a comprehensive map of the cell including genome sequence and gene expression data, information on protein localization, structure, function and expression, post-translational modifications, molecular and genetic interactions and phenotypic descriptions. Some of this broad set of functional genomics data has been already assembled for the budding yeast. Even though molecular cartography of the yeast cell is still far from comprehensive, functional genomics has begun to forge connections between disparate cellular events and to foster numerous hypotheses. Here we review several different genomics and proteomics technologies and describe bioinformatics methods for exploring these data to make new discoveries.

Chromosome Mapping↗
PubMed2003

Ac'septin' a signal: kinase regulation by septins.

Budding yeast monitor shape and the assembly of cytoskeletal structures and convey this information to regulators of cell division, but the molecular mechanisms responsible for monitoring and interpreting spatial information about the cytoskeleton remain poorly understood. A paper in the September issue of Molecular Cell shows that direct binding of components of the septin cytoskeleton may relieve autoinhibition of a conserved checkpoint kinase, creating a simple molecular device for sensing septin cytoskeleton organization.

Animals↗
PubMed2003

The global transcriptional activator of Saccharomyces cerevisiae, Gcr1p, mediates the response to glucose by stimulating protein synthesis and CLN-dependent cell cycle progression.

Growth of Saccharomyces cerevisiae requires coordination of cell cycle events (e.g., new cell wall deposition) with constitutive functions like energy generation and duplication of protein mass. The latter processes are stimulated by the phosphoprotein Gcr1p, a transcriptional activator that operates through two different Rap1p-mediated mechanisms to boost expression of glycolytic and ribosomal protein genes, respectively. Simultaneous disruption of both mechanisms results in a loss of glucose responsiveness and a dramatic drop in translation rate. Since a critical rate of protein synthesis (CRPS) is known to mediate passage through Start and determine cell size by modulating levels of Cln3p, we hypothesized that GCR1 regulates cell cycle progression by coordinating it with growth. We therefore constructed and analyzed gcr1delta cln3delta and gcr1delta cln1delta cln2delta strains. Both strains are temperature and cold sensitive; interestingly, they exhibit different arrest phenotypes. The gcr1delta cln3delta strain becomes predominantly unbudded with 1N DNA content (G1 arrest), whereas gcr1delta cln1delta cln2delta cells exhibit severe elongation and apparent M phase arrest. Further analysis demonstrated that the Rap1p/Gcr1p complex mediates rapid growth in glucose by stimulating both cellular metabolism and CLN transcription.

Base Sequence↗
PubMed2003

G1 transcription factors are differentially regulated in Saccharomyces cerevisiae by the Swi6-binding protein Stb1.

Stage-specific transcriptional programs are an integral feature of cell cycle regulation. In the budding yeast Saccharomyces cerevisiae, over 120 genes are coordinately induced in late G(1) phase by two heterodimeric transcription factors called SBF and MBF. Activation of SBF and MBF is an upstream initiator of key cell cycle events, including budding and DNA replication. SBF and MBF regulation is complex and genetically redundant, and the precise mechanism of G(1) transcriptional activation is unclear. Assays using SBF- and MBF-specific reporter genes revealed that the STB1 gene specifically affected MBF-dependent transcription. STB1 encodes a known Swi6-binding protein, but an MBF-specific function had not been previously suspected. Consistent with a specific role in regulating MBF, a STB1 deletion strain requires SBF for viability and microarray studies show a decrease in MBF-regulated transcripts in a swi4Delta mutant following depletion of Stb1. Chromatin immunoprecipitation experiments confirm that Stb1 localizes to promoters of MBF-regulated genes. Our data indicate that, contrary to previous models, MBF and SBF have unique components and might be distinctly regulated.

CDC28 Protein Kinase, S cerevisiae↗
PubMed2002

High-resolution genetic mapping with ordered arrays of Saccharomyces cerevisiae deletion mutants.

We present a method for high-resolution genetic mapping that takes advantage of the ordered set of viable gene deletion mutants, which form a set of colinear markers covering almost every centimorgan of the Saccharomyces cerevisiae genome, and of the synthetic genetic array (SGA) system, which automates the construction of double mutants formed by mating and meiotic recombination. The Cbk1 kinase signaling pathway, which consists minimally of CBK1, MOB2, KIC1, HYM1, and TAO3 (PAG1), controls polarized morphogenesis and activation of the Ace2 transcription factor. Deletion mutations in the Cbk1 pathway genes are tolerated differently by common laboratory strains of S. cerevisiae, being viable in the W303 background but dead in the S288C background. Genetic analysis indicated that the lethality of Cbk1 pathway deletions in the S288C background was suppressed by a single allele specific to the W303 background. SGA mapping (SGAM) was used to locate this W303-specific suppressor to the SSD1 locus, which contains a known polymorphism that appears to compromise SSD1 function. This procedure should map any mutation, dominant or recessive, whose phenotype is epistatic to wild type, that is, a phenotype that can be scored from a mixed population of cells obtained by germination of both mutant and wild-type spores. In principle, SGAM should be applicable to the analysis of multigenic traits. Large-scale construction of ordered mutations in other model organisms would broaden the application of this approach.

Adaptor Proteins, Signal Transducing↗
PubMed2002

Dissection of a complex phenotype by functional genomics reveals roles for the yeast cyclin-dependent protein kinase Pho85 in stress adaptation and cell integrity.

Cyclin-dependent kinases (Cdks) are key regulators of the cell division cycle. Pho85 is a multifunctional Cdk in budding yeast involved in aspects of metabolism, the cell cycle, cell polarity, and gene expression. Consistent with a broad spectrum of functions, Pho85 associates with a family of 10 cyclins and deletion of PHO85 causes a pleiotropic phenotype. Discovering the physiological substrates of protein kinases is a major challenge, and we have pursued a number of genomics approaches to reveal the processes regulated by Pho85 and to understand the root cause of reduced cellular fitness in pho85Delta mutant strains. We used a functional-genomics approach called synthetic genetic array (SGA) analysis to systematically identify strain backgrounds in which PHO85 is required for viability. In parallel, we used DNA microarrays to examine the genome-wide transcriptional consequences of deleting PHO85 or members of the Pho85 cyclin family. Using this pairwise approach coupled with phenotypic tests, we uncovered clear roles for Pho85 in cell integrity and the response to adverse growth conditions. Importantly, our combined approach allowed us to ascribe new aspects of the complex pho85 phenotype to particular cyclins; our data highlight a cell integrity function for the Pcl1,2 subgroup of Pho85 Cdks that is independent of a role for the Pho80-Pho85 kinase in the response to stress. Using a modification of the SGA technique to screen for suppressors of pho85Delta strain growth defects, we found that deletion of putative vacuole protein gene VTC4 suppressed the sensitivity of the pho85Delta strain to elevated CaCl(2) and many other stress conditions. Expression of VTC4 is regulated by Pho4, a transcription factor that is inhibited by the Pho80-Pho85 kinase. Genetic tests and electron microscopy experiments suggest that VTC4 is a key target of Pho4 and that Pho80-Pho85-mediated regulation of VTC4 expression is required for proper vacuole function and for yeast cell survival under a variety of suboptimal conditions. The integration of multiple genomics approaches is likely to be a generally useful strategy for extracting functional information from pleiotropic mutant phenotypes.

Adaptation, Physiological↗