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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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Tyr1P presents a specific pattern of phosphorylation along genes compared to Pol II.(A) Genome-wide profiling of Pol II (N20) and CTD isoforms (as in Figure 2) for different classes of binding levels indicate a distribution of Tyr1P more prominent at promoters vs gene bodies as compared to Pol II and Ser7P, but comparable to that of Ser5P. The indicated signal rank of the values is over an area encompassing TSS, GB, and 3′ ends of genes as indicated in the ‘Materials and methods–Gene selection and average binding profiles’. Note that more Tyr1P signal is found at 3′ ends as compared to Ser5P. (B) Spearman correlation plots of significantly enriched areas for Pol II and phospho-isoforms (genes size >2 kb) indicate that Tyr1P relates more to Pol II and early transcription marks at promoters than it does at gene bodies or 3′ends. Mean values for Spearman correlation were computed at [TSS-500 bp;TSS+500 bp], [TSS+1000 bp; 3′end-500 bp], and [3′end-500 bp; 3′end+1000 bp] (‘Materials and methods–Correlation of biological replicates and cross-correlation’).DOI:http://dx.doi.org/10.7554/eLife.02105.012
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fig2s6: Tyr1P presents a specific pattern of phosphorylation along genes compared to Pol II.(A) Genome-wide profiling of Pol II (N20) and CTD isoforms (as in Figure 2) for different classes of binding levels indicate a distribution of Tyr1P more prominent at promoters vs gene bodies as compared to Pol II and Ser7P, but comparable to that of Ser5P. The indicated signal rank of the values is over an area encompassing TSS, GB, and 3′ ends of genes as indicated in the ‘Materials and methods–Gene selection and average binding profiles’. Note that more Tyr1P signal is found at 3′ ends as compared to Ser5P. (B) Spearman correlation plots of significantly enriched areas for Pol II and phospho-isoforms (genes size >2 kb) indicate that Tyr1P relates more to Pol II and early transcription marks at promoters than it does at gene bodies or 3′ends. Mean values for Spearman correlation were computed at [TSS-500 bp;TSS+500 bp], [TSS+1000 bp; 3′end-500 bp], and [3′end-500 bp; 3′end+1000 bp] (‘Materials and methods–Correlation of biological replicates and cross-correlation’).DOI:http://dx.doi.org/10.7554/eLife.02105.012

Mentions: To assess its relation to transcription genome-wide, we next performed Tyr1P ChIP-seq, using 3D12 mAb, and compared it to Pol II and the other phospho-isoforms. We isolated significantly associated regions based on the signal distribution of the background-subtracted data (Figure 2—figure supplement 1B) and found that Pol II and all isoforms, including Tyr1P, correlated with transcription levels of genes (Figure 2—figure supplement 2). At many gene locations, a predominant signal of Tyr1P at promoters was observed (Figure 2B, Figure 2—figure supplement 3). We further confirmed this by genome-wide profiling of Pol II isoforms at coding-gene locations (Figure 2C, Figure 2—figure supplement 4, Figure 2—figure supplement 5A for Ser2P profile). Our profiling analysis shows that Tyr1P signal is predominantly found at promoters similarly to Ser5P, weak or essentially absent at gene bodies and weak at 3′ends in contrast to Ser2P elongating mark and Ser7P (associated to both promoters and gene bodies). These observations are further supported by quantification of signals at various genic sections (Figure 2—figure supplement 6) and reinforce our conclusion that human Tyr1P is mainly associated to promoters in an early, post-initiation step of transcription. Although we did not further investigate this possibility, in the accompanying manuscript, Hsin et al show that Chicken Tyr1 is found phosphorylated in the nucleoplasm, raising the possibility that Tyr1P is also associated with recruitment of the enzyme and transcription initiation.


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)

Tyr1P presents a specific pattern of phosphorylation along genes compared to Pol II.(A) Genome-wide profiling of Pol II (N20) and CTD isoforms (as in Figure 2) for different classes of binding levels indicate a distribution of Tyr1P more prominent at promoters vs gene bodies as compared to Pol II and Ser7P, but comparable to that of Ser5P. The indicated signal rank of the values is over an area encompassing TSS, GB, and 3′ ends of genes as indicated in the ‘Materials and methods–Gene selection and average binding profiles’. Note that more Tyr1P signal is found at 3′ ends as compared to Ser5P. (B) Spearman correlation plots of significantly enriched areas for Pol II and phospho-isoforms (genes size >2 kb) indicate that Tyr1P relates more to Pol II and early transcription marks at promoters than it does at gene bodies or 3′ends. Mean values for Spearman correlation were computed at [TSS-500 bp;TSS+500 bp], [TSS+1000 bp; 3′end-500 bp], and [3′end-500 bp; 3′end+1000 bp] (‘Materials and methods–Correlation of biological replicates and cross-correlation’).DOI:http://dx.doi.org/10.7554/eLife.02105.012
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Related In: Results  -  Collection

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fig2s6: Tyr1P presents a specific pattern of phosphorylation along genes compared to Pol II.(A) Genome-wide profiling of Pol II (N20) and CTD isoforms (as in Figure 2) for different classes of binding levels indicate a distribution of Tyr1P more prominent at promoters vs gene bodies as compared to Pol II and Ser7P, but comparable to that of Ser5P. The indicated signal rank of the values is over an area encompassing TSS, GB, and 3′ ends of genes as indicated in the ‘Materials and methods–Gene selection and average binding profiles’. Note that more Tyr1P signal is found at 3′ ends as compared to Ser5P. (B) Spearman correlation plots of significantly enriched areas for Pol II and phospho-isoforms (genes size >2 kb) indicate that Tyr1P relates more to Pol II and early transcription marks at promoters than it does at gene bodies or 3′ends. Mean values for Spearman correlation were computed at [TSS-500 bp;TSS+500 bp], [TSS+1000 bp; 3′end-500 bp], and [3′end-500 bp; 3′end+1000 bp] (‘Materials and methods–Correlation of biological replicates and cross-correlation’).DOI:http://dx.doi.org/10.7554/eLife.02105.012
Mentions: To assess its relation to transcription genome-wide, we next performed Tyr1P ChIP-seq, using 3D12 mAb, and compared it to Pol II and the other phospho-isoforms. We isolated significantly associated regions based on the signal distribution of the background-subtracted data (Figure 2—figure supplement 1B) and found that Pol II and all isoforms, including Tyr1P, correlated with transcription levels of genes (Figure 2—figure supplement 2). At many gene locations, a predominant signal of Tyr1P at promoters was observed (Figure 2B, Figure 2—figure supplement 3). We further confirmed this by genome-wide profiling of Pol II isoforms at coding-gene locations (Figure 2C, Figure 2—figure supplement 4, Figure 2—figure supplement 5A for Ser2P profile). Our profiling analysis shows that Tyr1P signal is predominantly found at promoters similarly to Ser5P, weak or essentially absent at gene bodies and weak at 3′ends in contrast to Ser2P elongating mark and Ser7P (associated to both promoters and gene bodies). These observations are further supported by quantification of signals at various genic sections (Figure 2—figure supplement 6) and reinforce our conclusion that human Tyr1P is mainly associated to promoters in an early, post-initiation step of transcription. Although we did not further investigate this possibility, in the accompanying manuscript, Hsin et al show that Chicken Tyr1 is found phosphorylated in the nucleoplasm, raising the possibility that Tyr1P is also associated with recruitment of the enzyme and transcription initiation.

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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