Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Nuclear Localization of Hog1 MAPK is Not Necessary for Resistance to Hyperosmotic Stress

ABSTRACT: In budding yeast, this signaling pathway— the high-osmolarity glycerol (HOG) response —culminates in dual phosphorylation and nuclear translocation of the MAPK, Hog1 (ortholog of mammalian p38/SAPK). Induction of at least 50 genes requires nuclear Hog1, implying that transcriptional up-regulation is necessary to cope with hyperosmotic stress. Contrary to this expectation, we found that cells lacking the karyopherin (Nmd5) required for Hog1 nuclear import or in which Hog1 was permanently anchored at the plasma membrane(HOG1-CCAAX) (or both) withstood hyperosmotic challenge by three different solutes (1 M sorbitol, KCl or NaCl). In cells where activated Hog1 is excluded from the nucleus, there was little change in transcriptional program after exposure to hyperosmotic shock (comparable to hog1∆ cells), as judged by examining several diagnostic mRNAs and by global transcript measurements using microarrays. Systematic genetic analysis ruled out the need for any transcription factor known to be influenced by Hog1 (Hot1, Msn2, Msn4, Sko1 and Smp1). Keywords: Time course of stress response gene expression array The transcriptomes' of HOG1-GFP, hog1del, and HOG1-CCAAX strains before and after 60 min hyperosmotic shock with 1M sorbitol at 25C were compared. Three biological replicates were done, with the first biological replicate done in technical triplicate, and the final two biological replicates beind done in technical duplicate by in slide duplication of features. Several supplementary files attached to the Series are summarized below: GSE8703 B1, B2, B3 Tscombined_matrix files show the averages of the technical replicates for each biological replicate. GSE8703 B1, B2, B3 norm_to_0_matrix files show the fold change over the time course (log2 time 60 - log2 time 0) for each of the three strains in each biological replicate. GSE8703 Figure_S6 shows the 30 genes with the most significant difference between the HOG1-GFP strain and the hog1del strain as determined by SAM. Genes were identified as: >3 fold induction in HOG1-GFP over 60 minute hyperosmotic shock and <3 fold induction in hog1del over 60 minute hyperosmotic shock.

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Jesse Patterson

PROVIDER: E-GEOD-8703 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Stress resistance and signal fidelity independent of nuclear MAPK function.

Westfall Patrick J PJ Patterson Jesse C JC Chen Raymond E RE Thorner Jeremy J

Proceedings of the National Academy of Sciences of the United States of America 20080821 34

Elevated external solute stimulates a conserved MAPK cascade that elicits responses that maintain osmotic balance. The yeast high-osmolarity glycerol (HOG) pathway activates Hog1 MAPK (mammalian ortholog p38alpha/SAPKalpha), which enters the nucleus and induces expression of >50 genes, implying that transcriptional up-regulation is necessary to cope with hyperosmotic stress. Contrary to this expectation, we show here that cells lacking the karyopherin required for Hog1 nuclear import or in which ...[more]

PMID: 18719124

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Project description:Oral streptococci, including Streptococcus gordonii, and Actinomyces naeslundii, are consistently found to be the most abundant bacteria in the early stages of dental plaque accumulation. These organisms interact physically (coaggregate) in vitro and in vivo. We hypothesized that coaggregation between S. gordonii and A. naeslundii leads to changes in gene expression in the partner organisms. Furthermore, we predicted that coaggregation-induced changes in phenotype contribute to the success of streptococci and actinomyces in dental plaque. To assess the responses of S. gordonii to coaggregation with A. naeslundii, RNA was extracted from S. gordonii cells 3 h after inducing coaggregation with A. naeslundii or from equivalent S. gordonii monocultures. The two RNA populations were reverse transcribed and compared by competitive hybridization with an S. gordonii genomic microarray. The most striking feature of the response to coaggregation was a profound change in expression of S. gordonii genes involved in arginine biosynthesis and transport. Subsequent experiments demonstrated that coaggregation with A. naeslundii stabilizes arginine biosynthesis in S. gordonii and enables growth under low-arginine conditions, such as those present in human saliva. Keywords: Cell-cell interaction The S. gordonii microarrays consist of 2195 70-mer oligonucleotides representing 2151 open reading frames, each repeated six times on the array. Chemically defined medium (CDM), was based in Tereleckyj’s FMC with minor modifications (Jakubovics et al., 2008). For coaggregate cultures, concentrated suspensions of S. gordonii DL1 (Challis) and A. naeslundii MG1 in CDM were mixed, vortexed and diluted to 1 x 108 cfu/ml. Monocultures were set up identically, except that A. naeslundii cells were omitted. Cultures were incubated at 37oC for 3 h prior to harvesting and extraction of total RNA. Purified RNA was reverse transcribed and cDNAs were labelled with Cy3 or Cy5 dye. cDNAs from coaggregate cultures and from S. gordonii monocultures were competitively hybridized with the S. gordonii microarray. Three independent sets of cultures were used, and flip dye pairs were included for two of the biological replicates (ie 5 hybridizations in total). In control experiments, cDNA derived from A.naeslundii monocultures did not hybridize with the S. gordonii microarrays. Data represent the ratios of gene expression in coaggregated S. gordonii compared with S. gordonii monocultured cells.

Dataset Information

Nuclear Localization of Hog1 MAPK is Not Necessary for Resistance to Hyperosmotic Stress

Publications

Stress resistance and signal fidelity independent of nuclear MAPK function.

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