Interferon-γ-dependent tyrosine phosphorylation of MEKK4 via Pyk2 is regulated by annexin II and SHP2 in keratinocytes

Ursula M. Halfter, Zachary E. Derbyshire, Richard R. Vaillancourt

Research output: Contribution to journalArticlepeer-review

30 Scopus citations

Abstract

IFNγ (interferon-γ) binding to its cognate receptor results, through JAK (Janus kinase), in direct activation of receptor-bound STAT1 (signal transducer and activator of transcription 1), although there is evidence for additional activation of a MAPK (mitogen-activated protein kinase) pathway. In the present paper, we report IFNγ-dependent activation of the MEKK4 (MAPK/extracellular-signal-regulated kinase kinase kinase 4) pathway in HaCaT human keratinocytes. MEKK4 is tyrosine-phosphorylated and the IFNγ-dependent phosphorylation requires intracellular calcium. Calcium-dependent phosphorylation of MEKK4 is mediated by Pyk2. Moreover, MEKK4 and Pyk2 co-localize in an IFNγ-dependent manner in the perinuclear region. Furthermore, the calcium-binding protein, annexin II, and the calcium-regulated kinase. Pyk2, co-immunoprecipitate with MEKK4 after treatment with IFNγ. Immunofluorescence imaging of HaCaT cells shows an IFNγ-dependent co-localization of annexin II with Pyk2 in the perinuclear region, suggesting that annexin II mediates the calcium-dependent regulation of Pyk2. Tyrosine phosphorylation of MEKK4 correlates with its activity to phosphorylate MKK6 (MAPK kinase 6) in vitro and subsequent p38 MAPK activation in an IFNγ-dependent manner. Additional studies demonstrate that the SH2 (Src homology 2)-domain-containing tyrosine phosphatase SHP2 co-immunoprecipitates with MEKK4 in an IFNγ-dependent manner and co-localizes with MEKK4 after IFNγ stimulation in the perinuclear region in HaCaT cells. Furthermore, we provide evidence that SHP2 dephosphorylates MEKK4 and Pyk2, terminating the MEKK4-dependent branch of the IFNγ signalling pathway.

Original languageEnglish (US)
Pages (from-to)17-28
Number of pages12
JournalBiochemical Journal
Volume388
Issue number1
DOIs
StatePublished - May 15 2005

Keywords

  • Annexin II
  • Interferon-γ (IFNγ)
  • Mitogen-activated protein kinase/extracellular-signal-regulated kinase kinase kinase 4 (MEKK4)
  • Pyk2
  • Src-homology-2-domain-containing phosphatase (SHP2)

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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