1. Biochemistry and Chemical Biology
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A secretory kinase complex regulates extracellular protein phosphorylation

  1. Jixin Cui
  2. Junyu Xiao
  3. Vincent S Tagliabracci
  4. Jianzhong Wen
  5. Meghdad Rahdar
  6. Jack E Dixon  Is a corresponding author
  1. University of California, San Diego, United States
  2. Peking University, China
  3. Merck & Co, United States
  4. ISIS Pharmaceuticals, United States
Research Article
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Cite this article as: eLife 2015;4:e06120 doi: 10.7554/eLife.06120

Abstract

Although numerous extracellular phosphoproteins have been identified, the protein kinases within the secretory pathway have only recently been discovered, and their regulation is virtually unexplored. Fam20C is the physiological Golgi casein kinase, which phosphorylates many secreted proteins and is critical for proper biomineralization. Fam20A, a Fam20C paralog, is essential for enamel formation, but the biochemical function of Fam20A is unknown. Here we show that Fam20A potentiates Fam20C kinase activity and promotes the phosphorylation of enamel matrix proteins in vitro and in cells. Mechanistically, Fam20A is a pseudokinase that forms a functional complex with Fam20C, and this complex enhances extracellular protein phosphorylation within the secretory pathway. Our findings shed light on the molecular mechanism by which Fam20C and Fam20A collaborate to control enamel formation, and provide the first insight into the regulation of secretory pathway phosphorylation.

Article and author information

Author details

  1. Jixin Cui

    Department of Pharmacology, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  2. Junyu Xiao

    State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.

  3. Vincent S Tagliabracci

    Department of Pharmacology, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Jianzhong Wen

    Discovery Bioanalytics, Merck & Co, Rahway, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Meghdad Rahdar

    ISIS Pharmaceuticals, Carlsbad, United States
    Competing interests
    The authors declare that no competing interests exist.

  6. Jack E Dixon

    Department of Pharmacology, University of California, San Diego, San Diego, United States
    For correspondence
    jedixon@ucsd.edu
    Competing interests
    The authors declare that no competing interests exist.

Ethics

Animal experimentation: Procedures involving mice were reviewed and approved by the Institutional Animal Care and Use Committee (IACUC) at the UC San Diego (Protocol #S03039).

Reviewing Editor

  1. Tony Hunter, Salk Institute, United States

Publication history

  1. Received: December 16, 2014
  2. Accepted: March 18, 2015
  3. Accepted Manuscript published: March 19, 2015 (version 1)
  4. Version of Record published: May 6, 2015 (version 2)

Copyright

© 2015, Cui et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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Further reading

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics

    Structure of Fam20A reveals a pseudokinase featuring a unique disulfide pattern and inverted ATP-binding

    Jixin Cui et al.
    Research Advance Updated

    Mutations in FAM20A cause tooth enamel defects known as Amelogenesis Imperfecta (AI) and renal calcification. We previously showed that Fam20A is a secretory pathway pseudokinase and allosterically activates the physiological casein kinase Fam20C to phosphorylate secreted proteins important for biomineralization (Cui et al., 2015). Here we report the nucleotide-free and ATP-bound structures of Fam20A. Fam20A exhibits a distinct disulfide bond pattern mediated by a unique insertion region. Loss of this insertion due to abnormal mRNA splicing interferes with the structure and function of Fam20A, resulting in AI. Fam20A binds ATP in the absence of divalent cations, and strikingly, ATP is bound in an inverted orientation compared to other kinases. Fam20A forms a dimer in the crystal, and residues in the dimer interface are critical for Fam20C activation. Together, these results provide structural insights into the function of Fam20A and shed light on the mechanism by which Fam20A mutations cause disease.

    1. Biochemistry and Chemical Biology

    Protein Kinases: What is the point of pseudokinases?

    Saravanan Raju, Andrey S Shaw
    Insight

    The interaction between an active kinase and an ‘inactive’ pseudokinase provides clues about how these enzymes were regulated in the past, and how this regulation has evolved.

    1. Biochemistry and Chemical Biology

    A novel DNA primase-helicase pair encoded by SCCmec elements

    Aleksandra Bebel et al.
    Research Article

    Mobile genetic elements (MGEs) are a rich source of new enzymes, and conversely, understanding the activities of MGE-encoded proteins can elucidate MGE function. Here we biochemically characterize 3 proteins encoded by a conserved operon carried by the Staphylococcal Cassette Chromosome (SCCmec), an MGE that confers methicillin resistance to Staphylococcus aureus, creating MRSA strains. The first of these proteins, CCPol, is an active A-family DNA polymerase. The middle protein, MP, binds tightly to CCPol and confers upon it the ability to synthesize DNA primers de novo. The CCPol-MP complex is therefore a unique primase-polymerase enzyme unrelated to either known primase family. The third protein, Cch2, is a 3'-to-5' helicase. Cch2 additionally binds specifically to a dsDNA sequence downstream of its gene that is also a preferred initiation site for priming by CCPol-MP. Taken together, our results suggest that this is a functional replication module for SCCmec.