Arabidopsis thaliana
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ABSTRACT:
PROVIDER: PRJNA138167 | ENA |
REPOSITORIES: ENA
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Project description:Serine phosphorylation of conserved Y1S2P3T4S5P6S7 repeats of RNA polymerase II carboxy-terminal domain (RNAPII CTD) plays a central role in the regulation of transcription and co-transcriptional RNA processing. Maintenance of CTD phosphoserine-7 mark in Arabidopsis requires the CDKF;1 kinase, which mediates in vivo activation of downstream-acting CDKD CTD kinase family. Knockout mutations of CDKF;1 lead to over 50% reduction of RNAPII CTD Ser-7 phosphorylation as early as 7 days after germination in seedlings. The transcript profiling experiment aimed at determining how early changes in CTD Ser-7 phosphorylation affect global regulation of transcription. We used Affymetrix ATH1-121501 Genome Array to compare global transcript levels in wild type and cdkf;1-2 mutant seedlings 7 days after germination.
2011-09-01 | GSE27687 | GEO
Project description:Serine phosphorylation of conserved Y1S2P3T4S5P6S7 repeats of RNA polymerase II carboxy-terminal domain (RNAPII CTD) plays a central role in the regulation of transcription and co-transcriptional RNA processing. Maintenance of CTD phosphoserine-7 mark in Arabidopsis requires the CDKF;1 kinase, which mediates in vivo activation of downstream-acting CDKD CTD kinase family. Knockout mutations of CDKF;1 lead to over 50% reduction of RNAPII CTD Ser-7 phosphorylation as early as 7 days after germination in seedlings. The transcript profiling experiment aimed at determining how early changes in CTD Ser-7 phosphorylation affect global regulation of transcription. We used Affymetrix ATH1-121501 Genome Array to compare global transcript levels in wild type and cdkf;1-2 mutant seedlings 7 days after germination. Wild type and cdkf;1-2 mutant Arabidopsis (Col-0) seedlings were grown in solid MSAR medium (Koncz, C., Martini, N., Szabados, L., Hrouda, M., Bachmair, A., and Schell, J. (1994). Specialized vectors for gene tagging and expression studies. In: Plant Molecular Biology Manual, Gelvin,S., and Schilperoort, B. (eds.), Kluwer Academic Publishers, Dordrecht-Boston-London, B2, 1-22.) containing ½ concentration of macro elements and 0.5% sucrose in Petri-dishes at 22°C, 120 mol/m2s light intensity and 8h light/16h dark period. Seedling were collected at 4h after the start of the light period in liquid nitrogen for RNA preparation.
2011-08-31 | E-GEOD-27687 | biostudies-arrayexpress
Project description:Dynamic post-translational modification of RNA polymerase II (RNAPII) coordinates the co-transcriptional recruitment of enzymatic complexes that regulate chromatin states and co-transcriptional processing of nascent RNA. Extensive phosphorylation of serine residues occurs at the structurally-disordered C-terminal domain (CTD) of the largest RNAPII subunit, which is composed of multiple heptapeptide repeats with consensus sequence Y1-S2-P3-T4-S5-P6-S7. Serine-5 and Serine-7 phosphorylation mark transcription initiation, whereas Serine-2 phosphorylation coincides with productive elongation. In vertebrates, the CTD has eight non-canonical substitutions of Serine-7 into Lysine-7, which can be acetylated (K7ac). Here, we describe for the first time mono- and di-methylation of CTD Lysine-7 residues (K7me1 and K7me2). K7me1 and K7me2 are observed during the earliest transcription stages and precede or accompany Serine-5 and Serine-7 phosphorylation. Genome wide mapping of 2 novel RNAPII post-translational modifications (CTD-K7me1 and CTD-K7me2) in mouse ES cells.
2015-12-22 | E-GEOD-72876 | biostudies-arrayexpress
Project description:Dynamic post-translational modification of RNA polymerase II (RNAPII) coordinates the co-transcriptional recruitment of enzymatic complexes that regulate chromatin states and co-transcriptional processing of nascent RNA. Extensive phosphorylation of serine residues occurs at the structurally-disordered C-terminal domain (CTD) of the largest RNAPII subunit, which is composed of multiple heptapeptide repeats with consensus sequence Y1-S2-P3-T4-S5-P6-S7. Serine-5 and Serine-7 phosphorylation mark transcription initiation, whereas Serine-2 phosphorylation coincides with productive elongation. In vertebrates, the CTD has eight non-canonical substitutions of Serine-7 into Lysine-7, which can be acetylated (K7ac). Here, we describe for the first time mono- and di-methylation of CTD Lysine-7 residues (K7me1 and K7me2). K7me1 and K7me2 are observed during the earliest transcription stages and precede or accompany Serine-5 and Serine-7 phosphorylation.
2015-12-22 | GSE72876 | GEO
Project description:DYRK1A is a dosage-sensitive protein kinase that fulfills key roles during development and in tissue homeostasis, and its dysregulation results in human pathologies. DYRK1A is present in both the nucleus and cytoplasm of mammalian cells, although its nuclear function remains unclear. Genome-wide analysis of DYRK1A-associated loci reveals that the kinase is recruited preferentially to promoters of genes actively transcribed by RNA polymerase II (RNAPII), which are functionally associated with translation, RNA processing and cell cycle. DYRK1A-bound promoter sequences are highly enriched in a conserved palindromic motif, which is necessary to drive DYRK1A-dependent transcriptional activation. DYRK1A phosphorylates the carboxy-terminal domain (CTD) of RNAPII at Ser2 and Ser5. Depletion of DYRK1A results in reduced association of RNAPII at the target promoters as well as hypophosphorylation of the CTD of RNAPII along the target gene bodies. Accordingly, we propose that DYRK1A acts as a transcriptional regulator by acting as a novel CTD kinase. Occupancy of the kinase DYRK1A in two different cell lines and in two different growing conditions.
2015-03-11 | E-GEOD-63712 | biostudies-arrayexpress
Project description:Release of promoter-proximal paused RNA polymerase II (Pol II) during early elongation is a critical step in transcriptional regulation in metazoan cells. Paused Pol II release is thought to require the kinase activity of cyclin-dependent kinase 9 (CDK9) for the phosphorylation of DRB sensitivity-inducing factor, negative elongation factor, and C-terminal domain (CTD) serine-2 of Pol II. We found that Pol II-associated factor 1 (PAF1) is a critical regulator of paused Pol II release, that positive transcription elongation factor b (P-TEFb) directly regulates the initial recruitment of PAF1 complex (PAF1C) to genes, and that the subsequent recruitment of CDK12 is dependent on PAF1C. These findings reveal cooperativity among P-TEFb, PAF1C, and CDK12 in pausing release and Pol II CTD phosphorylation. Comparison of the chromatin occupancy of [1] PAF1, CDC73, LEO1, CTR9, total Pol II, and CTD serine 2-phosphorylated Pol II by ChIP-seq in THP1 cells; [2] PAF1, Pol II, Pol II (ser-5p), CDK12, and CDK9 by ChIP-seq in control and PAF1 knockdown cells; [3] LEO1 and Pol II by ChIP-seq in control and flavopiridol treated THP1 cells.
2015-12-11 | E-GEOD-62171 | biostudies-arrayexpress
Project description:RNA polymerase II (Pol II) was immunoprecipitated from Arabidopsis thaliana seedlings, using antibodies that specifically recognize: 1) C’ terminal Domain (CTD), 2) CTD Serine 5 Phosphorylation and 3) CTD Serine 2 Phosphorylation. Proteins that co-immunoprecipitate with different Pol II pools were analysed.
2018-10-17 | PXD006004 | Pride
Project description:The RNA polymerase II (RNApII) C-terminal domain (CTD)-interacting domain (CID) proteins are involved in two distinct termination pathways and recognize different phosphorylated forms of CTD. To investigate the role of differential CTDM-^VCID interactions in the choice of termination pathway, we altered the CTD-binding specificity of Nrd1 by domain swapping. ChIP-chip was performed to examine the effect of Nrd1 CID swapping on genome-wide RNA polymerase II (Rpb3 antibody, Neoclone) occupancy. Nrd1 with the CID from Rtt103 (Nrd1[CID-Rtt103]; strain YSB2445) causes read-through transcription at many genes, but can trigger termination where multiple Nrd1/Nab3-binding sites and serine 2 phosphorylated CTD co-exist.
2013-12-04 | E-MTAB-1626 | biostudies-arrayexpress
Project description:Spt6 is a multifunctional histone chaperone involved in the maintenance of chromatin structure during elongation by RNA polymerase II (Pol II). Spt6 has a tandem SH2 (tSH2) domain within its C-terminus that recognizes Pol II CTD peptides phosphorylated on Ser2, Ser5 or Try1 in vitro. Deleting the tSH2 domain, however, only has a partial effect on Spt6 occupancy in vivo, suggesting that more complex mechanisms are involved in the Spt6 recruitment. Our results show that the Ser2 kinases Bur1 and Ctk1, but not the Ser5 kinase Kin28, cooperate in recruiting Spt6, genome-wide. Interestingly, the Ser2 kinases promote the association of Spt6 in early transcribed regions and not toward the 3' end of genes, where phosphorylated Ser2 reaches its maximum level. Additionally, our results uncover an unexpected role for histone deacetylases (Rpd3 and Hos2) in promoting Spt6 interaction with elongating Pol II. Finally, our data suggest that phosphorylation of the Pol II CTD on Tyr1 promotes the association of Spt6 with the 3' end of transcribed genes, independently of Ser2 phosphorylation. Collectively, our results show that a complex network of interactions, involving the Spt6 tSH2 domain, CTD phosphorylation and histone deacetylases, coordinate the recruitment of Spt6 to transcribed genes in vivo. We examined the genome-wide distribution (using ChIP-chip) of Spt6. Spt6 occupancy was also assayed in mutants for CTD Serine 2 and Serine 5 kinases and in mutants for histone deacetylases. ChIPs were performed with a Myc-tagged version of Spt6. Most ChIPs (in Cy5) were hybridyzed against a control ChIP sample from an isogenic non-tagged strain (in Cy3). In the ChIP experiments with the spt6-202del mutant, non immunoprecipitated DNA (input) was used as the control. In addition to Spt6 ChIPs, the project includes RNAPII (Rpb3) ChIP-chip datasets, where an anti-Rpb3 antibody was used to ChIP RNAPII and non immunoprecipitated DNA (input) was used as the control. All ChIP-chip experiments were done in duplicates. Each microarray was normalized using the Lima Loess and replicates were combined using a weighted average method as previously described (Pokholok et al., 2005).
2014-09-25 | E-GEOD-61713 | biostudies-arrayexpress
Project description:The Carboxy-terminal domain (CTD) of RNA Polymerase II (RNAPII) in mammals undergoes extensive post-translational modification, which is essential for transcriptional initiation and elongation. Here, we show that the CTD of RNAPII is methylated at a single arginine (R1810) by the transcriptional co-activator CARM1. Although methylation at R1810 is present on the hyper-phosphorylated form of RNAPII in vivo, Ser-2 or Ser-5 phosphorylation inhibit CARM1 activity towards this site in vitro, suggesting that methylation occurs before transcription initiation. Mutation of R1810 results in the mis-expression of a variety of snRNAs and snoRNAs, an effect that is also observed in Carm1-/- MEFs. These results demonstrate that CTD methylation facilitates the expression of select RNAs, perhaps serving to discriminate the RNAPII-associated machinery recruited to distinct gene types. To address the function of RNAPII methylation, we generated Raji cell lines expressing an RNA Polymerase II resistant to α-amanitin and carrying either wild-type R1810 or an arginine to alanine substitution at that same residue, abolishing R1810 methylation of the CTD. In cells cultured in α-amanitin, the α-amanitin-resistant mutants fully replaced the functions of endogenous RNAPII, allowing us to study if gene-expression is affected by the absence of R1810me
2011-04-01 | E-GEOD-27315 | biostudies-arrayexpress