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A NETWORK OF PROTEIN COMPLEXES SIGNALING TO THE RNA POLYMERASE II TRANSCRIPTION APPARATUS IN HUMAN CELLS
Author(s) -
Coulombe Benoit,
Jeronimo Célia,
Bouchard Annie,
Thérien Cynthia,
Chua Gordon,
Bergeron Dominique,
Bourassa Sylvie,
Greenblatt Jack,
Chabot Benoit,
Poirier Guy,
Hughes Timothy R.,
Forget Diane
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a945-a
Subject(s) - rna polymerase ii , rna splicing , microbiology and biotechnology , biology , chromatin , gene silencing , transcription (linguistics) , messenger rna , gene expression , rna polymerase , sr protein , alternative splicing , rna binding protein , computational biology , rna , gene , promoter , genetics , linguistics , philosophy
The enzyme RNA polymerase (RNAP) II, whose activity is modulated by many regulatory proteins, synthesizes all mRNA in eukaryotes. To explore the network of protein complexes that signal to RNAP II in mammalian cells, we have developed a systematic procedure where the expression of affinity tagged polypeptides and the purification of their associated proteins, as they prospectively exist in vivo, is coupled to protein identification by mass spectrometry (MS). All newly detected interactions were validated experimentally and novel interaction partners were reciprocally tagged, purified and the eluates characterized by MS. Thirty tagged polypeptides yield a network of more than 325 validated interactions. Our results identify nine previously uncharacterized proteins to which we have inferred a function on the basis of their network connections, bioinformatics and the effect of silencing their homologues on global gene expression in yeast. These polypeptides participate in defining at least two novel interfaces linking RNAP II to specific regulatory machineries, one involved in alternative pre‐mRNA splicing and arginine methylation and the other, comprised of chaperone/scaffolding‐related factors, linking RNAP II to the Mediator, the Integrator and chromatin‐remodeling factors. Together, our results define novel transcriptional regulators, pathways and mechanisms.