Browsing by Subject "RNA-POLYMERASE-II"

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  • Liu, Xiaonan; Salokas, Kari; Tamene, Fitsum; Jiu, Yaming; Weldatsadik, Rigbe G.; öhman, Tiina; Varjosalo, Markku (2018)
    Protein-protein interactions govern almost all cellular functions. These complex networks of stable and transient associations can be mapped by affinity purification mass spectrometry (AP-MS) and complementary proximity-based labeling methods such as BioID. To exploit the advantages of both strategies, we here design and optimize an integrated approach combining AP-MS and BioID in a single construct, which we term MAC-tag. We systematically apply the MAC-tag approach to 18 subcellular and 3 sub-organelle localization markers, generating a molecular context database, which can be used to define a protein's molecular location. In addition, we show that combining the AP-MS and BioID results makes it possible to obtain interaction distances within a protein complex. Taken together, our integrated strategy enables the comprehensive mapping of the physical and functional interactions of proteins, defining their molecular context and improving our understanding of the cellular interactome.
  • Virtanen, Jori A.; Vartiainen, Maria K. (2017)
    In addition to its essential roles as part of the cytoskeleton, actin has also been linked to many processes in the nucleus. Recent data has demonstrated the presence of both monomeric and polymeric actin in the nucleus, and implied distinct functional roles for these actin pools. Monomeric actin seems to be involved in regulation of gene expression through transcription factors, chromatin regulating complexes and RNA polymerases. In addition to cytoplasmic actin regulators, nuclear proteins, such as emerin, can regulate actin polymerization properties specifically in this compartment. Besides of structural roles, nuclear actin filaments may be required for organizing the nuclear contents and for the maintenance of genomic integrity.
  • Beimforde, Christina; Feldberg, Kathrin; Nylinder, Stephan; Rikkinen, Jouko; Tuovila, Hanna; Doerfelt, Heinrich; Gube, Matthias; Jackson, Daniel J.; Reitner, Joachim; Seyfullah, Leyla J.; Schmidt, Alexander R. (2014)
  • Ovaska, Kristian; Matarese, Filomena; Grote, Korbinian; Charapitsa, Iryna; Cervera, Alejandra; Liu, Chengyu; Reid, George; Seifert, Martin; Stunnenberg, Hendrik G.; Hautaniemi, Sampsa (2013)
  • Taube, Ran; Peterlin, Boris Matija (2013)
  • Ekumi, Kingsley; Paculova, Hana; Lenasi, Tina; Pospichalova, Vendula; Boesken, Christian A.; Rybarikova, Jana; Bryja, Vitezslav; Geyer, Matthias; Blazek, Dalibor; Barboric, Matjaz (2015)
    The Cdk12/CycK complex promotes expression of a subset of RNA polymerase II genes, including those of the DNA damage response. CDK12 is among only nine genes with recurrent somatic mutations in high-grade serous ovarian carcinoma. However, the influence of thesemutations on the Cdk12/CycK complex and their link to cancerogenesis remain ill-defined. Here, we show that most mutations prevent formation of the Cdk12/CycK complex, rendering the kinase inactive. By examining the mutations within the Cdk12/CycK structure, we find that they likely provoke structural rearrangements detrimental to Cdk12 activation. Our mRNA expression analysis of the patient samples containing the CDK12 mutations reveals coordinated downregulation of genes critical to the homologous recombination DNA repair pathway. Moreover, we establish that the Cdk12/CycK complex occupies these genes and promotes phosphorylation of RNA polymerase II at Ser2. Accordingly, we demonstrate that the mutant Cdk12 proteins fail to stimulate the faithful DNA double strand break repair via homologous recombination. Together, we provide the molecular basis of how mutated CDK12 ceases to function in ovarian carcinoma. We propose that CDK12 is a tumor suppressor of which the loss-of-function mutations may elicit defects in multiple DNA repair pathways, leading to genomic instability underlying the genesis of the cancer.
  • Bugai, Andrii; Quaresma, Alexandre J. C.; Friedel, Caroline C.; Lenasi, Tina; Düster, Robert; Sibley, Christopher R.; Fuginaga, Koh; Kukanja, Petra; Hennig, Thomas; Blasius, Melanie; Geyer, Matthias; Ule, Jernej; Dölken, Lars; Barboric, Matjaz (2019)
    DNA damage response (DDR) involves dramatic transcriptional alterations, the mechanisms of which remain ill defined. Here, we show that following genotoxic stress, the RNA-binding motif protein 7 (RBM7) stimulates RNA polymerase II (Pol II) transcription and promotes cell viability by activating the positive transcription elongation factor b (P-TEFb) via its release from the inhibitory 7SK small nuclear ribonucleoprotein (7SK snRNP). This is mediated by activation of p38MAPK, which triggers enhanced binding of RBM7 with core subunits of 7SK snRNP. In turn, P-TEFb relocates to chromatin to induce transcription of short units, including key DDR genes and multiple classes of non-coding RNAs. Critically, interfering with the axis of RBM7 and P-TEFb provokes cellular hypersensitivity to DNA-damage-inducing agents due to activation of apoptosis. Our work uncovers the importance of stress-dependent stimulation of Pol II pause release, which enables a pro-survival transcriptional response that is crucial for cell fate upon genotoxic insult.
  • Zumer, Kristina; Plemenitas, Ana; Saksela, Kalle; Peterlin, B. Matija (2011)