title: Critical role of novel Thr-219 autophosphorylation for the cellular function of PKCtheta in T lymphocytes.
abstract: Phosphopeptide mapping identified a major autophosphorylation site, phospho (p)Thr-219, between the tandem C1 domains of the regulatory fragment in protein kinase C (PKC)theta. Confirmation of this identification was derived using (p)Thr-219 antisera that reacted with endogenous PKCtheta in primary CD3+ T cells after stimulation with phorbol ester, anti-CD3 or vanadate. The T219A mutation abrogated the capacity of PKCtheta to mediate NF-kappaB, NF-AT and interleukin-2 promoter transactivation, and reduced PKCtheta's ability in Jurkat T cells to phosphorylate endogenous cellular substrates. In particular, the T219A mutation impaired crosstalk of PKCtheta with Akt/PKBalpha in NF-kappaB activation. Yet, this novel (p)Thr-219 site did not affect catalytic activity or second-messenger lipid-binding activity in vitro. Instead, the T219A mutation prevented proper recruitment of PKCtheta in activated T cells. The PKCthetaT219A mutant defects were largely rescued by addition of a myristoylation signal to force its proper membrane localization. We conclude that autophosphorylation of PKCtheta at Thr-219 plays an important role in the correct targeting and cellular function of PKCtheta upon antigen receptor ligation.
authors.affiliation: Department for Medical Genetics, Molecular and Clinical Pharmacology, Innsbruck Medical University, Innsbruck, Austria.
authors.forename: Nikolaus
authors.initials: N
authors.lastname: Thuille
authors.forename: Isabelle
authors.initials: I
authors.lastname: Heit
authors.forename: Friedrich
authors.initials: F
authors.lastname: Fresser
authors.forename: Nina
authors.initials: N
authors.lastname: Krumböck
authors.forename: Birgit
authors.initials: B
authors.lastname: Bauer
authors.forename: Sabine
authors.initials: S
authors.lastname: Leuthaeusser
authors.forename: Sascha
authors.initials: S
authors.lastname: Dammeier
authors.forename: Caroline
authors.initials: C
authors.lastname: Graham
authors.forename: Terry D
authors.initials: TD
authors.lastname: Copeland
authors.forename: Steve
authors.initials: S
authors.lastname: Shaw
authors.forename: Gottfried
authors.initials: G
authors.lastname: Baier
mesh_terms: D002462:Cell Membrane / Q000201:enzymology; D004789:Enzyme Activation; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D019169:Jurkat Cells; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D000074121:Protein Kinase C-theta; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D013601:T-Lymphocytes / Q000201:enzymology*; D013912:Threonine / Q000378:metabolism*; D015533:Transcriptional Activation
chemical_list: D007376:Interleukin-2; D007527:Isoenzymes; D013912:Threonine; D051057:Proto-Oncogene Proteins c-akt; C076233:PRKCQ protein, human; D011493:Protein Kinase C; D000074121:Protein Kinase C-theta