title: Identification of a binding site in c-Ab1 tyrosine kinase for the C-terminal repeated domain of RNA polymerase II.
abstract: The c-abl proto-oncogene encodes a nuclear tyrosine kinase that can phosphorylate the mammalian RNA polymerase II (RNAP II) on its C-terminal repeated domain (CTD) in vitro. Phosphorylation of the CTD has previously been shown to require the tyrosine kinase and the SH2 domain of Abl. We show here that a CTD-interacting domain (CTD-ID) at the C-terminal region of c-Abl is also required. Deletion of the CTD-ID causes the Km 0.4 microM to increase by 2 orders of magnitude. Direct binding of the CTD-ID to CTD and to RNAP II could be demonstrated in vitro. Phosphorylation of a recombinant glutathione S-transferase-CTD by c-Abl was observed in cotransfected COS cells. Mutant Abl proteins lacking the CTD-ID, while capable of autophosphorylation, neither phosphorylated nor associated with the glutathione S-transferase-CTD in vivo. Transient overexpression of c-Abl also led to a four- to fivefold increase in the phosphotyrosine content of the RNAP II large subunit. Moreover, the large subunit of RNAP II could be coprecipitated with c-Abl. Tyrosine phosphorylation of the coprecipitated RNAP II was again dependent on the presence of the CTD-ID in Abl. Finally, the ability of c-Abl to phosphorylate and associate with RNAP II could be correlated with the enhancement of transcription by c-Abl in transient cotransfection assays. Taken together, these observations demonstrate that c-Abl can function as a CTD kinase in vitro as well as in vivo.
authors.affiliation: Department of Biology and Center for Molecular Genetics, University of California, San Diego, La Jolla, California 93093-0347, USA.
authors.forename: R
authors.initials: R
authors.lastname: Baskaran
authors.forename: G G
authors.initials: GG
authors.lastname: Chiang
authors.forename: J Y
authors.initials: JY
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D001859:Bone Neoplasms; D002460:Cell Line; D002522:Chlorocebus aethiops; D016384:Consensus Sequence; D016313:Genes, abl; D005982:Glutathione Transferase; D006801:Humans; D007668:Kidney; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D012516:Osteosarcoma; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000032:analysis; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D000090063:Proto-Oncogene Mas; D016315:Proto-Oncogene Proteins c-abl / Q000378:metabolism*; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D017384:Sequence Deletion; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D011993:Recombinant Fusion Proteins; D019000:Phosphotyrosine; D005982:Glutathione Transferase; D011505:Protein-Tyrosine Kinases; D016315:Proto-Oncogene Proteins c-abl; D012319:RNA Polymerase II