High-Throughput Genetic and Gene Expression Analysis of the RNAPII-CTD Reveals Unexpected Connections to SRB10/CDK8

English version České info

The C-terminal domain (CTD) of RNA polymerase II (RNAPII) is composed of heptapeptide repeats, which play a key regulatory role in gene expression. Using genetic interaction, chromatin immunoprecipitation followed by microarrays (ChIP-on-chip) and mRNA expression analysis, we found that truncating the CTD resulted in distinct changes to cellular function. Truncating the CTD altered RNAPII occupancy, leading to not only decreases, but also increases in mRNA levels. The latter were largely mediated by promoter elements and in part were linked to the transcription factor Rpn4. The mediator subunit Cdk8 was enriched at promoters of these genes, and its removal not only restored normal mRNA and RNAPII occupancy levels, but also reduced the abnormally high cellular amounts of Rpn4. This suggested a positive role of Cdk8 in relationship to RNAPII, which contrasted with the observed negative role at the activated INO1 gene. Here, loss of CDK8 suppressed the reduced mRNA expression and RNAPII occupancy levels of CTD truncation mutants.

Vyšlo v časopise: High-Throughput Genetic and Gene Expression Analysis of the RNAPII-CTD Reveals Unexpected Connections to SRB10/CDK8. PLoS Genet 9(8): e32767. doi:10.1371/journal.pgen.1003758
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.pgen.1003758

Souhrn

Zdroje

1. AllisonLA, MoyleM, ShalesM, InglesCJ (1985) Extensive homology among the largest subunits of eukaryotic and prokaryotic RNA polymerases. Cell 42 : 599–610.

2. CordenJL, CadenaDL, AhearnJMJr, DahmusME (1985) A unique structure at the carboxyl terminus of the largest subunit of eukaryotic RNA polymerase II. Proc Natl Acad Sci U S A 82 : 7934–7938.

3. HeidemannM, HintermairC, VossK, EickD (2012) Dynamic phosphorylation patterns of RNA polymerase II CTD during transcription. Biochim Biophys Acta 1829 : 55–62.

4. NonetM, SweetserD, YoungRA (1987) Functional redundancy and structural polymorphism in the large subunit of RNA polymerase II. Cell 50 : 909–915.

5. AllisonLA, WongJK, FitzpatrickVD, MoyleM, InglesCJ (1988) The C-terminal domain of the largest subunit of RNA polymerase II of saccharomyces cerevisiae, drosophila melanogaster, and mammals: A conserved structure with an essential function. Mol Cell Biol 8 : 321–329.

6. ZehringWA, LeeJM, WeeksJR, JokerstRS, GreenleafAL (1988) The C-terminal repeat domain of RNA polymerase II largest subunit is essential in vivo but is not required for accurate transcription initiation in vitro. Proc Natl Acad Sci U S A 85 : 3698–3702.

7. ScafeC, ChaoD, LopesJ, HirschJP, HenryS, et al. (1990) RNA polymerase II C-terminal repeat influences response to transcriptional enhancer signals. Nature 347 : 491–4.

8. LuH, FloresO, WeinmannR, ReinbergD (1991) The nonphosphorylated form of RNA polymerase II preferentially associates with the preinitiation complex. Proc Natl Acad Sci U S A 88 : 10004–10008.

9. KoborMS, GreenblattJ (2002) Regulation of transcription elongation by phosphorylation. Biochim Biophys Acta 1577 : 261–275.

10. MaxT, SogaardM, SvejstrupJQ (2007) Hyperphosphorylation of the C-terminal repeat domain of RNA polymerase II facilitates dissociation of its complex with mediator. J Biol Chem 282 : 14113–14120.

11. McCrackenS, FongN, RosoninaE, YankulovK, BrothersG, et al. (1997) 5′-capping enzymes are targeted to pre-mRNA by binding to the phosphorylated carboxy-terminal domain of RNA polymerase II. Genes Dev 11 : 3306–3318.

12. KimM, SuhH, ChoEJ, BuratowskiS (2009) Phosphorylation of the yeast Rpb1 C-terminal domain at serines 2, 5, and 7. J Biol Chem 284 : 26421–26426.

13. AkhtarMS, HeidemannM, TietjenJR, ZhangDW, ChapmanRD, et al. (2009) TFIIH kinase places bivalent marks on the carboxy-terminal domain of RNA polymerase II. Mol Cell 34 : 387–393.

14. ChoEJ, KoborMS, KimM, GreenblattJ, BuratowskiS (2001) Opposing effects of Ctk1 kinase and Fcp1 phosphatase at ser 2 of the RNA polymerase II C-terminal domain. Genes Dev 15 : 3319–3329.

15. MayerA, HeidemannM, LidschreiberM, SchreieckA, SunM, et al. (2012) CTD tyrosine phosphorylation impairs termination factor recruitment to RNA polymerase II. Science 336 : 1723–1725.

16. KoborMS, ArchambaultJ, LesterW, HolstegeFC, GileadiO, et al. (1999) An unusual eukaryotic protein phosphatase required for transcription by RNA polymerase II and CTD dephosphorylation in S. cerevisiae. Mol Cell 4 : 55–62.

17. ChoH, KimTK, ManceboH, LaneWS, FloresO, et al. (1999) A protein phosphatase functions to recycle RNA polymerase II. Genes Dev 13 : 1540–1552.

18. ZhangDW, MosleyAL, RamisettySR, Rodriguez-MolinaJB, WashburnMP, et al. (2012) Ssu72 phosphatase-dependent erasure of phospho-Ser7 marks on the RNA polymerase II C-terminal domain is essential for viability and transcription termination. J Biol Chem 287 : 8541–8551.

19. NonetML, YoungRA (1989) Intragenic and extragenic suppressors of mutations in the heptapeptide repeat domain of saccharomyces cerevisiae RNA polymerase II. Genetics 123 : 715–24.

20. ThompsonCM, KoleskeAJ, ChaoDM, YoungRA (1993) A multisubunit complex associated with the RNA polymerase II CTD and TATA-binding protein in yeast. Cell 73 : 1361–1375.

21. RobinsonPJ, BushnellDA, TrnkaMJ, BurlingameAL, KornbergRD (2012) Structure of the mediator head module bound to the carboxy-terminal domain of RNA polymerase II. Proc Natl Acad Sci U S A 109 : 17931–17935.

22. AndrauJC, van de PaschL, LijnzaadP, BijmaT, KoerkampMG, et al. (2006) Genome-wide location of the coactivator mediator: Binding without activation and transient Cdk8 interaction on DNA. Mol Cell 22 : 179–192.

23. ZhuX, WirenM, SinhaI, RasmussenNN, LinderT, et al. (2006) Genome-wide occupancy profile of mediator and the Srb8-11 module reveals interactions with coding regions. Mol Cell 22 : 169–178.

24. GuglielmiB, van BerkumNL, KlapholzB, BijmaT, BoubeM, et al. (2004) A high resolution protein interaction map of the yeast mediator complex. Nucleic Acids Res 32 : 5379–5391.

25. BhoiteLT, YuY, StillmanDJ (2001) The Swi5 activator recruits the mediator complex to the HO promoter without RNA polymerase II. Genes Dev 15 : 2457–2469.

26. HanSJ, LeeYC, GimBS, RyuGH, ParkSJ, et al. (1999) Activator-specific requirement of yeast mediator proteins for RNA polymerase II transcriptional activation. Mol Cell Biol 19 : 979–988.

27. GalbraithMD, DonnerAJ, EspinosaJM (2010) CDK8: A positive regulator of transcription. Transcription 1 : 4–12.

28. TsaiKL, SatoS, Tomomori-SatoC, ConawayRC, ConawayJW, et al. (2013) A conserved mediator-CDK8 kinase module association regulates mediator-RNA polymerase II interaction. Nat Struct Mol Biol 20 : 611–619.

29. HengartnerCJ, MyerVE, LiaoSM, WilsonCJ, KohSS, et al. (1998) Temporal regulation of RNA polymerase II by Srb10 and Kin28 cyclin-dependent kinases. Mol Cell 2 : 43–53.

30. LiuY, KungC, FishburnJ, AnsariAZ, ShokatKM, et al. (2004) Two cyclin-dependent kinases promote RNA polymerase II transcription and formation of the scaffold complex. Mol Cell Biol 24 : 1721–35.

31. HirstM, KoborMS, KuriakoseN, GreenblattJ, SadowskiI (1999) GAL4 is regulated by the RNA polymerase II holoenzyme-associated cyclin-dependent protein kinase SRB10/CDK8. Mol Cell 3 : 673–8.

32. CollinsSR, RoguevA, KroganNJ (2010) Quantitative genetic interaction mapping using the E-MAP approach. Methods Enzymol 470 : 205–231.

33. WestML, CordenJL (1995) Construction and analysis of yeast RNA polymerase II CTD deletion and substitution mutations. Genetics 140 : 1223–1233.

34. HolstegeFC, JenningsEG, WyrickJJ, LeeTI, HengartnerCJ, et al. (1998) Dissecting the regulatory circuitry of a eukaryotic genome. Cell 95 : 717–728.

35. MayerA, LidschreiberM, SiebertM, LeikeK, SodingJ, et al. (2010) Uniform transitions of the general RNA polymerase II transcription complex. Nat Struct Mol Biol 17 : 1272–1278.

36. SchulzeJM, HentrichT, NakanishiS, GuptaA, EmberlyE, et al. (2011) Splitting the task: Ubp8 and Ubp10 deubiquitinate different cellular pools of H2BK123. Genes Dev 25 : 2242–2247.

37. TietjenJR, ZhangDW, Rodriguez-MolinaJB, WhiteBE, AkhtarMS, et al. (2010) Chemical-genomic dissection of the CTD code. Nat Struct Mol Biol 17 : 1154–1161.

38. KimH, EricksonB, LuoW, SewardD, GraberJH, et al. (2010) Gene-specific RNA polymerase II phosphorylation and the CTD code. Nat Struct Mol Biol 17 : 1279–1286.

39. BatailleAR, JeronimoC, JacquesPE, LarameeL, FortinME, et al. (2012) A universal RNA polymerase II CTD cycle is orchestrated by complex interplays between kinase, phosphatase, and isomerase enzymes along genes. Mol Cell 45 : 158–170.

40. PokholokDK, HarbisonCT, LevineS, ColeM, HannettNM, et al. (2005) Genome-wide map of nucleosome acetylation and methylation in yeast. Cell 122 : 517–527.

41. XiaoT, HallH, KizerKO, ShibataY, HallMC, et al. (2003) Phosphorylation of RNA polymerase II CTD regulates H3 methylation in yeast. Genes Dev 17 : 654–663.

42. SuhMH, MeyerPA, GuM, YeP, ZhangM, et al. (2010) A dual interface determines the recognition of RNA polymerase II by RNA capping enzyme. J Biol Chem 285 : 34027–34038.

43. ChapmanRD, HeidemannM, AlbertTK, MailhammerR, FlatleyA, et al. (2007) Transcribing RNA polymerase II is phosphorylated at CTD residue serine-7. Science 318 : 1780–2.

44. FuchsSM, KizerKO, BrabergH, KroganNJ, StrahlBD (2011) RNA polymerase II carboxy-terminal domain phosphorylation regulates protein stability of the Set2 methyltransferase and histone H3 di -⁠ and trimethylation at lysine 36. J Biol Chem 287 : 3249–3256.

45. BricknerD, CajigasI, Fondufe-MittendorfY, AhmedS, LeeP, et al. (2007) H2A.Z-mediated localization of genes at the nuclear periphery confers epigenetic memory of previous transcriptional state. PLoS Biol 5: e81.

46. FanX, StruhlK (2009) Where does mediator bind in vivo? PLoS One 4: e5029.

47. van de PeppelJ, KettelarijN, van BakelH, KockelkornTT, van LeenenD, et al. (2005) Mediator expression profiling epistasis reveals a signal transduction pathway with antagonistic submodules and highly specific downstream targets. Mol Cell 19 : 511–522.

48. JuD, XuH, WangX, XieY (2007) Ubiquitin-mediated degradation of Rpn4 is controlled by a phosphorylation-dependent ubiquitylation signal. Biochim Biophys Acta 1773 : 1672–1680.

49. WongJMS, InglesCJ (2001) A compromised yeast RNA polymerase II enhances UV sensitivity in the absence of global genome nucleotide excision repair. Molecular and General Genetics 264 : 842–851.

50. KruegelU, RobisonB, DangeT, KahlertG, DelaneyJR, et al. (2011) Elevated proteasome capacity extends replicative lifespan in saccharomyces cerevisiae. PLoS Genet 7: e1002253.

51. WangX, XuH, HaSW, JuD, XieY (2010) Proteasomal degradation of Rpn4 in saccharomyces cerevisiae is critical for cell viability under stressed conditions. Genetics 184 : 335–342.

52. OwsianikG, BalzilL, GhislainM (2002) Control of 26S proteasome expression by transcription factors regulating multidrug resistance in saccharomyces cerevisiae. Mol Microbiol 43 : 1295–1308.

53. NelsonC, GotoS, LundK, HungW, SadowskiI (2003) Srb10/Cdk8 regulates yeast filamentous growth by phosphorylating the transcription factor Ste12. Nature 421 : 187–190.

54. RaithathaS, SuTC, LourencoP, GotoS, SadowskiI (2012) Cdk8 regulates stability of the transcription factor Phd1 to control pseudohyphal differentiation of saccharomyces cerevisiae. Mol Cell Biol 32 : 664–674.

55. HahnJS, NeefDW, ThieleDJ (2006) A stress regulatory network for co-ordinated activation of proteasome expression mediated by yeast heat shock transcription factor. Mol Microbiol 60 : 240–251.

56. RosoninaE, DuncanSM, ManleyJL (2012) Sumoylation of transcription factor Gcn4 facilitates its Srb10-mediated clearance from promoters in yeast. Genes Dev 26 : 350–355.

57. ChiY, HuddlestonMJ, ZhangX, YoungRA, AnnanRS, et al. (2001) Negative regulation of Gcn4 and Msn2 transcription factors by Srb10 cyclin-dependent kinase. Genes Dev 15 : 1078–1092.

58. LipfordJR, SmithGT, ChiY, DeshaiesRJ (2005) A putative stimulatory role for activator turnover in gene expression. Nature 438 : 113–116.

59. LongtineMS, McKenzieA3rd, DemariniDJ, ShahNG, WachA, et al. (1998) Additional modules for versatile and economical PCR-based gene deletion and modification in saccharomyces cerevisiae. Yeast 14 : 953–961.

60. GuarenteL, PtashneM (1981) Fusion of escherichia coli lacZ to the cytochrome c gene of saccharomyces cerevisiae. Proc Natl Acad Sci U S A 78 : 2199–2203.

61. van WageningenS, KemmerenP, LijnzaadP, MargaritisT, BenschopJJ, et al. (2010) Functional overlap and regulatory links shape genetic interactions between signaling pathways. Cell 143 : 991–1004.

62. LenstraTL, BenschopJJ, KimT, SchulzeJM, BrabersNA, et al. (2011) The specificity and topology of chromatin interaction pathways in yeast. Mol Cell 42 : 536–549.

63. ChangDT, HuangCY, WuCY, WuWS (2011) YPA: An integrated repository of promoter features in saccharomyces cerevisiae. Nucleic Acids Res 39: D647–52.

64. Carlson M, Falcon S, Pages H, Li N. Org.sc.sgd.db: Genome wide annotation for yeast. R package version 2.7.1.

65. FalconS, GentlemanR (2007) Using GOstats to test gene lists for GO term association. Bioinformatics 23 : 257–258.

66. SchulzeJM, JacksonJ, NakanishiS, GardnerJM, HentrichT, et al. (2009) Linking cell cycle to histone modifications: SBF and H2B monoubiquitination machinery and cell-cycle regulation of H3K79 dimethylation. Mol Cell 35 : 626–641.

67. KeoghMC, BuratowskiS (2004) Using chromatin immunoprecipitation to map cotranscriptional mRNA processing in saccharomyces cerevisiae. Methods Mol Biol 257 : 1–16.

68. DroitA, CheungC, GottardoR (2010) rMAT–an R/Bioconductor package for analyzing ChIP-chip experiments. Bioinformatics 26 : 678–679.

69. HentrichT, SchulzeJM, EmberlyE, KoborMS (2012) CHROMATRA: A galaxy tool for visualizing genome-wide chromatin signatures. Bioinformatics 28 : 717–718.

70. GuarenteL (1983) Yeast promoters and lacZ fusions designed to study expression of cloned genes in yeast. Methods Enzymol 101 : 181–191.