Membrane-bound steel factor induces more persistent tyrosine kinase activation and longer life span of c-kit gene-encoded protein than its soluble form

Keisuke Miyazawa, David A. Williams, Akihiko Gotoh, Jiroh Nishimaki, Hal E. Broxmeyer, Keisuke Toyama

Research output: Contribution to journalArticlepeer-review

214 Scopus citations

Abstract

Alternative splicing of exon 6 results in the production of two isoforms of Steel factor (SLF): the membrane-bound and soluble forms. To investigate differences in the kinetics of c-kit tyrosine kinase activated by these two isoforms, we used a stromal cell line (Sl/Sl4) established from Sl/Sl homozygous murine embryo fetal liver and its stable transfectants containing either hSCF248 cDNA (including exon 6; secreted form) or hSCF220 cDNA (lacking exon 6; membrane-bound form) as the source of each isoform. interaction of factor dependent myeloid cell line MO7e with stromal cells producing either isoform resulted in activated c-kit tyrosine kinase and induction of the same series of tyrosine phosphorylated cellular proteins in MO7e cells. However, Sl4-h220 (membrane-bound form) induced more persistent activation of c-kit kinase than Sl4-h248 (soluble form) did. Flow cytometric analysis and pulse-chase studies using [35S]methionine showed that Sl4- h248 induced rapid downmodulation of cell-surface c-kit expression and its protein degradation in MO7e cells, whereas Sl4-h220 induced more prolonged life span of c-kit protein. Addition of soluble recombinant human SLF to Sl4-h220 cultures enhanced reduction of cell-surface c-kit expression and its protein degradation. Because the kinetics of c-kit inactivation strikingly fits with the protein degradation rates of c-kit under the conditions described above, rapid proteolysis of c-kit protein induced by soluble SLF stimulation may function as a 'turn-off switch' for activated c- kit kinase.

Original languageEnglish (US)
Pages (from-to)641-649
Number of pages9
JournalBlood
Volume85
Issue number3
DOIs
StatePublished - Feb 1 1995

ASJC Scopus subject areas

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology

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