Characterization of the tyrosine kinase Tnk1 and its binding with phospholipase C-γ1

Donna M. Felschow, Curt I. Civin, Gerard T. Hoehn

Research output: Contribution to journalArticle

Abstract

Tnk1 is a nonreceptor tyrosine kinase cloned from CD34+/Lin-/CD38- hematopoietic stem/progenitor cells. The cDNA predicts a 72-kDa protein containing an NH2-terminal kinase, a Src Homology 3 (SH3) domain, and a proline-rich (PR) tail. We generated rabbit antiserum to a GST-Tnk1 (SH3) fusion protein. Affinity-purified anti-Tnk1 antibodies specifically recognized a 72-kDa protein in Tnk1-transfected COS-1 cells and cells which express Tnk1 mRNA. Western blot analysis indicated that Tnk1 is expressed in fetal blood cells, but not in any other hematopoietic tissues examined. Tnk1 immunoprecipitated from cell lysates possessed kinase activity and was tyrosine phosphorylated. In binding experiments with a panel of GST-fusion constructs, only GST-PLC-γ1 (SH3) interacted with in vitro translated Tnk1. GST-protein precipitations from cell lysates confirmed that GST-PLC-γ1 (SH3) associated with endogenously expressed Tnk1. Conversely, GST-Tnk1 (PR) protein constructs complexed with endogenously expressed PLC-γ1. The association of Tnk1 with PLC-γ1 suggests a role for Tnk1 in phospholipid signal transduction. (C) 2000 Academic Press.

Original languageEnglish (US)
Pages (from-to)294-301
Number of pages8
JournalBiochemical and Biophysical Research Communications
Volume273
Issue number1
DOIs
StatePublished - Jun 24 2000

Keywords

  • Ack
  • PLC-γ1
  • SII3 domain
  • Tnk1
  • Tyrosine kinase

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

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