title: A Single Kinase Generates the Majority of the Secreted Phosphoproteome.
abstract: The existence of extracellular phosphoproteins has been acknowledged for over a century. However, research in this area has been undeveloped largely because the kinases that phosphorylate secreted proteins have escaped identification. Fam20C is a kinase that phosphorylates S-x-E/pS motifs on proteins in milk and in the extracellular matrix of bones and teeth. Here, we show that Fam20C generates the majority of the extracellular phosphoproteome. Using CRISPR/Cas9 genome editing, mass spectrometry, and biochemistry, we identify more than 100 secreted phosphoproteins as genuine Fam20C substrates. Further, we show that Fam20C exhibits broader substrate specificity than previously appreciated. Functional annotations of Fam20C substrates suggest roles for the kinase beyond biomineralization, including lipid homeostasis, wound healing, and cell migration and adhesion. Our results establish Fam20C as the major secretory pathway protein kinase and serve as a foundation for new areas of investigation into the role of secreted protein phosphorylation in human biology and disease.
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Vincent S
authors.initials: VS
authors.lastname: Tagliabracci
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Sandra E
authors.initials: SE
authors.lastname: Wiley
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA; Department of Chemistry, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Xiao
authors.initials: X
authors.lastname: Guo
authors.affiliation: Department of Biophysics, University of Texas, Southwestern Medical Center, Dallas, TX 75390-9050, USA.
authors.forename: Lisa N
authors.initials: LN
authors.lastname: Kinch
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Eric
authors.initials: E
authors.lastname: Durrant
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Jianzhong
authors.initials: J
authors.lastname: Wen
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Junyu
authors.initials: J
authors.lastname: Xiao
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Jixin
authors.initials: J
authors.lastname: Cui
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Kim B
authors.initials: KB
authors.lastname: Nguyen
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: James L
authors.initials: JL
authors.lastname: Engel
authors.affiliation: Department of Chemistry, University of Wisconsin-Madison, Madison, WI 53706, USA; Department of Biomolecular Chemistry, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Joshua J
authors.initials: JJ
authors.lastname: Coon
authors.affiliation: Department of Biophysics, University of Texas, Southwestern Medical Center, Dallas, TX 75390-9050, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
authors.forename: Nick
authors.initials: N
authors.lastname: Grishin
authors.affiliation: Department of Biomedical Sciences, University of Padova, 35121 Padova, Italy; Venetian Institute of Molecular Medicine, 35129 Padova, Italy.
authors.forename: Lorenzo A
authors.initials: LA
authors.lastname: Pinna
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: David J
authors.initials: DJ
authors.lastname: Pagliarini
authors.affiliation: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA; Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA; Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: jedixon@mail.ucsd.edu.
authors.forename: Jack E
authors.initials: JE
authors.lastname: Dixon
mesh_terms: D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000378:metabolism; D047389:Casein Kinase I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002448:Cell Adhesion; D002465:Cell Movement; D002556:Cerebrospinal Fluid Proteins / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055786:Gene Knockout Techniques; D063990:Gene Ontology; D006801:Humans; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000032:analysis; D055571:Secretory Pathway; D013379:Substrate Specificity
chemical_list: D001798:Blood Proteins; D002556:Cerebrospinal Fluid Proteins; D016326:Extracellular Matrix Proteins; D010750:Phosphoproteins; D047389:Casein Kinase I; C524449:FAM20C protein, human