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Tyrosine phosphorylation of RNA polymerase II CTD is associated with antisense promoter transcription and active enhancers in mammalian cells.

Descostes N, Heidemann M, Spinelli L, Schüller R, Maqbool MA, Fenouil R, Koch F, Innocenti C, Gut M, Gut I, Eick D, Andrau JC - Elife (2014)

Bottom Line: Post-translational modifications of the CTD coordinate the transcription cycle and various steps of mRNA maturation.Here we describe Tyr1 phosphorylation (Tyr1P) as a hallmark of promoter (5' associated) Pol II in mammalian cells, in contrast to what was described in yeast.Mutation of Tyr1 to phenylalanine (Y1F) prevents the formation of the hyper-phosphorylated Pol IIO form, induces degradation of Pol II to the truncated Pol IIB form, and results in a lethal phenotype.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie de Marseille-Luminy, Université Aix-Marseille, Marseille, France Centre National de la Recherche Scientifique (CNRS) UMR6102, Marseille, France Inserm U631, Marseille, France Institut de Génétique Moléculaire de Montpellier (IGMM), CNRS-UMR5535, Montpellier, France.

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Examples of Tyr1P enhancer association upstream or downstream of CXCR4, DUSP2, and IER5 genes.As in Figure 4, light orange and blue rectangles highlight enhancer and promoter locations with higher H3K4me3 at promoters and higher H3K4me1 at enhancers. Relative amount of Tyr1P is higher at enhancers as compared to Pol II and to promoters. H3K4me3 level at CXCR4 IGR was observed but is not visible due to the scale used and because of high level of signal at promoter.DOI:http://dx.doi.org/10.7554/eLife.02105.019
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fig4s2: Examples of Tyr1P enhancer association upstream or downstream of CXCR4, DUSP2, and IER5 genes.As in Figure 4, light orange and blue rectangles highlight enhancer and promoter locations with higher H3K4me3 at promoters and higher H3K4me1 at enhancers. Relative amount of Tyr1P is higher at enhancers as compared to Pol II and to promoters. H3K4me3 level at CXCR4 IGR was observed but is not visible due to the scale used and because of high level of signal at promoter.DOI:http://dx.doi.org/10.7554/eLife.02105.019

Mentions: Many groups including ours have shown that highly active and tissue-specific enhancers are transcribed by Pol II in various tissues (De Santa et al., 2010; Kim et al., 2010; Koch et al., 2011; Natoli and Andrau, 2012). These enhancers can also be hallmarked by the occurrence of H3K4me1high/H3K4me3low epigenetic marks combination (Koch et al., 2011; Pekowska et al., 2011). To investigate if Tyr1P can be detected at enhancers, we first isolated intergenic regions (IGRs) with stringent criteria in B-cells using Pol II, H3K4me1, and me3 signals. These were further discriminated from noncoding promoters using the relative ratio of H3K4me1/me3 (Pekowska et al., 2011; Li et al., 2012), and from both unannotated coding and some long intergenic noncoding genes using the absence of H3K36me3 that marks gene bodies (Guttman et al., 2009). Using these criteria, we isolated 390 B-cells enhancers (Figure 4—figure supplement 1A–D). Our selection was further validated using tissue-specificity analyses (Figure 4—figure supplement 1E) indicating IGRs associated with genes specific to B-cells. We next performed profiling of the various Pol II isoforms at these enhancers. As before (Koch et al., 2011), we observed that these IGRs were associated with Ser5P (Figure 4A) but not with Ser2P Pol II (not shown) as well as with short transcripts (reflecting paused transcription) and a discrete nucleosome depleted region. Consistent with early elongating Pol II at enhancers, we found signal for both Ser7P and Tyr1P at these IGRs. Importantly, Tyr1P appeared more bound to enhancers as compared to promoters and total Pol II (Figure 4B,C, Figure 4—figure supplement 2), suggesting that Tyr1 is more phosphorylated than Ser5 or Ser7 at enhancers and represent a hallmark of these essential areas of the genome. Additionally, Tyr1P also displayed the best correlation with Pol II at isolated enhancers (Figure 4D). Finally, using an independent selection for active enhancers based on H3K27ac brought very similar results (Figure 4—figure supplement 3). Together, our investigations showed that Tyr1P is a strong signature of Pol II-transcribed active enhancers associated with tissue-specific gene expression.10.7554/eLife.02105.017Figure 4.Tyr1P is a hallmark of enhancers relative to Pol II and promoters signal.


Tyrosine phosphorylation of RNA polymerase II CTD is associated with antisense promoter transcription and active enhancers in mammalian cells.

Descostes N, Heidemann M, Spinelli L, Schüller R, Maqbool MA, Fenouil R, Koch F, Innocenti C, Gut M, Gut I, Eick D, Andrau JC - Elife (2014)

Examples of Tyr1P enhancer association upstream or downstream of CXCR4, DUSP2, and IER5 genes.As in Figure 4, light orange and blue rectangles highlight enhancer and promoter locations with higher H3K4me3 at promoters and higher H3K4me1 at enhancers. Relative amount of Tyr1P is higher at enhancers as compared to Pol II and to promoters. H3K4me3 level at CXCR4 IGR was observed but is not visible due to the scale used and because of high level of signal at promoter.DOI:http://dx.doi.org/10.7554/eLife.02105.019
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4042876&req=5

fig4s2: Examples of Tyr1P enhancer association upstream or downstream of CXCR4, DUSP2, and IER5 genes.As in Figure 4, light orange and blue rectangles highlight enhancer and promoter locations with higher H3K4me3 at promoters and higher H3K4me1 at enhancers. Relative amount of Tyr1P is higher at enhancers as compared to Pol II and to promoters. H3K4me3 level at CXCR4 IGR was observed but is not visible due to the scale used and because of high level of signal at promoter.DOI:http://dx.doi.org/10.7554/eLife.02105.019
Mentions: Many groups including ours have shown that highly active and tissue-specific enhancers are transcribed by Pol II in various tissues (De Santa et al., 2010; Kim et al., 2010; Koch et al., 2011; Natoli and Andrau, 2012). These enhancers can also be hallmarked by the occurrence of H3K4me1high/H3K4me3low epigenetic marks combination (Koch et al., 2011; Pekowska et al., 2011). To investigate if Tyr1P can be detected at enhancers, we first isolated intergenic regions (IGRs) with stringent criteria in B-cells using Pol II, H3K4me1, and me3 signals. These were further discriminated from noncoding promoters using the relative ratio of H3K4me1/me3 (Pekowska et al., 2011; Li et al., 2012), and from both unannotated coding and some long intergenic noncoding genes using the absence of H3K36me3 that marks gene bodies (Guttman et al., 2009). Using these criteria, we isolated 390 B-cells enhancers (Figure 4—figure supplement 1A–D). Our selection was further validated using tissue-specificity analyses (Figure 4—figure supplement 1E) indicating IGRs associated with genes specific to B-cells. We next performed profiling of the various Pol II isoforms at these enhancers. As before (Koch et al., 2011), we observed that these IGRs were associated with Ser5P (Figure 4A) but not with Ser2P Pol II (not shown) as well as with short transcripts (reflecting paused transcription) and a discrete nucleosome depleted region. Consistent with early elongating Pol II at enhancers, we found signal for both Ser7P and Tyr1P at these IGRs. Importantly, Tyr1P appeared more bound to enhancers as compared to promoters and total Pol II (Figure 4B,C, Figure 4—figure supplement 2), suggesting that Tyr1 is more phosphorylated than Ser5 or Ser7 at enhancers and represent a hallmark of these essential areas of the genome. Additionally, Tyr1P also displayed the best correlation with Pol II at isolated enhancers (Figure 4D). Finally, using an independent selection for active enhancers based on H3K27ac brought very similar results (Figure 4—figure supplement 3). Together, our investigations showed that Tyr1P is a strong signature of Pol II-transcribed active enhancers associated with tissue-specific gene expression.10.7554/eLife.02105.017Figure 4.Tyr1P is a hallmark of enhancers relative to Pol II and promoters signal.

Bottom Line: Post-translational modifications of the CTD coordinate the transcription cycle and various steps of mRNA maturation.Here we describe Tyr1 phosphorylation (Tyr1P) as a hallmark of promoter (5' associated) Pol II in mammalian cells, in contrast to what was described in yeast.Mutation of Tyr1 to phenylalanine (Y1F) prevents the formation of the hyper-phosphorylated Pol IIO form, induces degradation of Pol II to the truncated Pol IIB form, and results in a lethal phenotype.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie de Marseille-Luminy, Université Aix-Marseille, Marseille, France Centre National de la Recherche Scientifique (CNRS) UMR6102, Marseille, France Inserm U631, Marseille, France Institut de Génétique Moléculaire de Montpellier (IGMM), CNRS-UMR5535, Montpellier, France.

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