Genome-wide location analysis of the stress-activated MAP kinase Hog1 in yeast

Harvard University, Cambridge, Massachusetts, United States
Methods (Impact Factor: 3.65). 12/2006; 40(3):272-8. DOI: 10.1016/j.ymeth.2006.06.007
Source: PubMed


MAP kinase signal transduction pathways play a critical role in eukaryotic cells to unleash complex transcriptional programs to properly adapt to changing environments. The MAP kinase Hog1 upon activation is physically recruited to the chromatin of osmostress responsive genes. This allowed us to use in vivo chromatin immunoprecipitation in combination with microarrays (ChIP-Chip) to identify the transcriptional targets of Hog1 at the genomic scale. The ChIP-Chip method described here revealed that the stress-activated MAP kinase gets recruited to most of the osmoinducible genes. Interestingly Hog1 associates with both the 5' upstream and the 3' downstream sequences of stress genes. We confirmed by targeted ChIP at several stress genes that the MAP kinase crosslinks all over the transcribed regions in all cases tested. Taken together the genome-wide location analysis reported here is a powerful approach to determine the genomic binding patterns of an activated MAP kinase and will be of great interest to analyze other SAPKs under different environmental conditions.

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Available from: Markus Proft
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    • "Genome-wide studies using ChIP and microarray analysis have been instrumental in uncovering the presence of Hog1 associated with a number of stress-responsive genes as well as its localization at both promoter and coding regions of stress-responsive genes. However, the number of genes uncovered by these approaches has been rather limited and never totaled more than 70 genes [15-17]. The relevance of Hog1 in gene expression and RNA Pol II recruitment suggested that the number of genes with Hog1 association could have been underestimated. "
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    ABSTRACT: Background Cells are subjected to dramatic changes of gene expression upon environmental changes. Stress causes a general down-regulation of gene expression together with the induction of a set of stress-responsive genes. The p38-related stress-activated protein kinase Hog1 is an important regulator of transcription upon osmostress in yeast. Results Genome-wide localization studies of RNA polymerase II (RNA Pol II) and Hog1 showed that stress induced major changes in RNA Pol II localization, with a shift toward stress-responsive genes relative to housekeeping genes. RNA Pol II relocalization required Hog1, which was also localized to stress-responsive loci. In addition to RNA Pol II-bound genes, Hog1 also localized to RNA polymerase III-bound genes, pointing to a wider role for Hog1 in transcriptional control than initially expected. Interestingly, an increasing association of Hog1 with stress-responsive genes was strongly correlated with chromatin remodeling and increased gene expression. Remarkably, MNase-Seq analysis showed that although chromatin structure was not significantly altered at a genome-wide level in response to stress, there was pronounced chromatin remodeling for those genes that displayed Hog1 association. Conclusion Hog1 serves to bypass the general down-regulation of gene expression that occurs in response to osmostress, and does so both by targeting RNA Pol II machinery and by inducing chromatin remodeling at stress-responsive loci.
    Full-text · Article · Nov 2012 · Genome biology
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    • "In addition to its role in the regulation of the Rtg1/3 subcellular localization, Hog1 is further required for Rtg1/3 chromatin binding and transcriptional activity. It is known that Hog1 becomes intimately linked with stress-responsive loci upon osmostress, and this binding is dependent on the presence of stress-mediating transcriptional activators (Alepuz et al., 2001, 2003; Pascual-Ahuir et al., 2006; Pokholok et al., 2006; Proft et al., 2006). Similarly, Hog1 is recruited by Rtg1 to the RTG-dependent promoters in response to osmostress. "
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    ABSTRACT: Cells modulate expression of nuclear genes in response to alterations in mitochondrial function, a response termed retrograde (RTG) regulation. In budding yeast, the RTG pathway relies on Rtg1 and Rtg3 basic helix-loop-helix leucine Zipper transcription factors. Exposure of yeast to external hyperosmolarity activates the Hog1 Stress-Activated Protein Kinase (SAPK), which is a key player in the regulation of gene expression upon stress. Several transcription factors, including Sko1, Hot1, the redundant Msn2 and Msn4, and Smp1, have been shown to be directly controlled by the Hog1 SAPK. The mechanisms by which Hog1 regulates their activity differ from one to another. Here, we show that Rtg1 and Rtg3 transcription factors are new targets of the Hog1 SAPK. In response to osmostress, RTG-dependent genes are induced in a Hog1-dependent manner and Hog1 is required for Rtg1/3 complex nuclear accumulation. In addition, Hog1 activity regulates Rtg1/3 binding to chromatin and transcriptional activity. Therefore, Hog1 modulates Rtg1/3 complex activity by multiple mechanisms in response to stress. Overall, our data suggest that Hog1, through activation of the RTG pathway, contributes to ensure mitochondrial function as part of the Hog1-mediated osmoadaptive response.
    Preview · Article · Sep 2012 · Molecular biology of the cell
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    • "We identified 28 ORFs that are enriched for Hog1 during osmotic shock (listed in File S1). These findings are in general agreement with previous work (Pascual-Ahuir et al. 2006; Pokholok et al. 2006; Figure S3), but by using a higher resolution technique, we are able to more clearly differentiate between Hog1 present in regulatory regions vs. coding regions. We find that Hog1 occupies the ORFs that show the highest increase in RNA Pol II occupancy upon stress (Figure 1C, red points), while Hog1 is not present in ORFs that show a decrease in RNA Pol II occupancy in response to stress or in the ORFs of HOG pathway-activated genes that show only modest induction upon stress. "
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    ABSTRACT: When challenged with osmotic shock, Saccharomyces cerevisiae induces hundreds of genes, despite a concurrent reduction in overall transcriptional capacity. The stress-responsive MAP kinase Hog1 activates expression of specific genes through interactions with chromatin remodeling enzymes, transcription factors, and RNA polymerase II. However, it is not clear whether Hog1 is involved more globally in modulating the cell's transcriptional program during stress, in addition to activating specific genes. Here we show that large-scale redistribution of RNA Pol II from housekeeping to stress genes requires Hog1. We demonstrate that decreased RNA Pol II occupancy is the default outcome for highly expressed genes upon stress and that Hog1 is partially required for this effect. We find that Hog1 and RNA Pol II colocalize to open reading frames that bypass global transcriptional repression. These activation targets are specified by promoter binding of two osmotic stress-responsive transcription factors. The combination of reduced global transcription with a gene-specific override mechanism allows cells to rapidly switch their transcriptional program in response to stress.
    Full-text · Article · Sep 2012 · G3-Genes Genomes Genetics
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