title: Intra-domain Cross-talk Regulates Serine-arginine Protein Kinase 1-dependent Phosphorylation and Splicing Function of Transformer 2β1.
abstract: Transformer 2β1 (Tra2β1) is a splicing effector protein composed of a core RNA recognition motif flanked by two arginine-serine-rich (RS) domains, RS1 and RS2. Although Tra2β1-dependent splicing is regulated by phosphorylation, very little is known about how protein kinases phosphorylate these two RS domains. We now show that the serine-arginine protein kinase-1 (SRPK1) is a regulator of Tra2β1 and promotes exon inclusion in the survival motor neuron gene 2 (SMN2). To understand how SRPK1 phosphorylates this splicing factor, we performed mass spectrometric and kinetic experiments. We found that SRPK1 specifically phosphorylates 21 serines in RS1, a process facilitated by a docking groove in the kinase domain. Although SRPK1 readily phosphorylates RS2 in a splice variant lacking the N-terminal RS domain (Tra2β3), RS1 blocks phosphorylation of these serines in the full-length Tra2β1. Thus, RS2 serves two new functions. First, RS2 positively regulates binding of the central RNA recognition motif to an exonic splicing enhancer sequence, a phenomenon reversed by SRPK1 phosphorylation on RS1. Second, RS2 enhances ligand exchange in the SRPK1 active site allowing highly efficient Tra2β1 phosphorylation. These studies demonstrate that SRPK1 is a regulator of Tra2β1 splicing function and that the individual RS domains engage in considerable cross-talk, assuming novel functions with regard to RNA binding, splicing, and SRPK1 catalysis.
authors.affiliation: From the Department of Pharmacology, University of California at San Diego, La Jolla, California 92093-0636 and.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Jamros
authors.affiliation: From the Department of Pharmacology, University of California at San Diego, La Jolla, California 92093-0636 and.
authors.forename: Brandon E
authors.initials: BE
authors.lastname: Aubol
authors.affiliation: From the Department of Pharmacology, University of California at San Diego, La Jolla, California 92093-0636 and.
authors.forename: Malik M
authors.initials: MM
authors.lastname: Keshwani
authors.affiliation: the Molecular and Cellular Biochemistry Department, University of Kentucky, Lexington, Kentucky 40536.
authors.forename: Zhaiyi
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: the Molecular and Cellular Biochemistry Department, University of Kentucky, Lexington, Kentucky 40536.
authors.forename: Stefan
authors.initials: S
authors.lastname: Stamm
authors.affiliation: From the Department of Pharmacology, University of California at San Diego, La Jolla, California 92093-0636 and j2adams@ucsd.edu.
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Adams
mesh_terms: D001665:Binding Sites; D005091:Exons; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D054730:Protein Interaction Domains and Motifs; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012313:RNA / Q000235:genetics / Q000378:metabolism; D012326:RNA Splicing; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000737:chemistry; D000068103:Serine-Arginine Splicing Factors; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D055540:Survival of Motor Neuron 2 Protein / Q000235:genetics / Q000378:metabolism
chemical_list: D009419:Nerve Tissue Proteins; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; C526477:SMN2 protein, human; D055540:Survival of Motor Neuron 2 Protein; C097083:TRA2B protein, human; D000068103:Serine-Arginine Splicing Factors; D012694:Serine; D012313:RNA; C087728:SRPK1 protein, human; D017346:Protein Serine-Threonine Kinases
keywords: RNA; alternative splicing; kinetics; mass spectrometry (MS); protein kinase; rapid quench