Quantitative analysis of dynamic protein interactions during transcription reveals a role for casein kinase II in Polymerase-associated Factor (PAF) complex phosphorylation and regulation of histone H2B monoubiquitylation

Lynn Glowczewski Bedard, Raghuvar Dronamraju, Jenny L. Kerschner, Gerald O. Hunter, Elizabeth De Vlieger Axley, Asha K. Boyd, Brian D. Strahl, Amber Mosley

Research output: Contribution to journalArticle

8 Scopus citations

Abstract

Using affinity purification MS approaches, we have identified a novel role for casein kinase II (CKII) in the modification of the polymerase associated factor complex (PAF-C). Our data indicate that the facilitates chromatin transcription complex (FACT) interacts with CKII and may facilitate PAF complex phosphorylation. Posttranslational modification analysis of affinity-isolated PAF-C shows extensive CKII phosphorylation of all five subunits of PAF-C, although CKII subunits were not detected as interacting partners. Consistent with this, recombinant CKII or FACT-associated CKII isolated from cells can phosphorylate PAF-C in vitro, whereas no intrinsic kinase activity was detected in PAF-C samples. Significantly, PAF-C purifications combined with stable isotope labeling in cells (SILAC) quantitation for PAF-C phosphorylation from wild-type and CKII temperature-sensitive strains (cka1Δ cka2-8) showed that PAF-C phosphorylation at consensus CKII sites is significantly reduced in cka1Δ cka2-8 strains. Consistent with a role of CKII in FACT and PAF-C function, we show that decreased CKII function in vivo results in decreased levels of histone H2B lysine 123 monoubiquitylation, a modification dependent on FACT and PAF-C. Taken together, our results define a coordinated role of CKII and FACT in the regulation of RNA polymerase II transcription through chromatin via phosphorylation of PAF-C.

Original languageEnglish (US)
Pages (from-to)13410-13420
Number of pages11
JournalJournal of Biological Chemistry
Volume291
Issue number26
DOIs
StatePublished - Jun 24 2016

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

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