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

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 (SI/SI4) established from SI/SI 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, SI4- h220 (membrane-bound form) induced more persistent activation of c-kit kinase than SI4-h248 (soluble form) did. Flow cytometric analysis and pulse-chase studies using [35S]methionine showed that SI4-h248 induced rapid downmodulation of cell-surface c-kit expression and its protein degradation in MO7e cells, whereas SI4-h220 induced more prolonged life span of c-kit protein. Addition of soluble recombinant human SLF to SI4- 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.

Diverse isoforms of colony‐stimulating factor‐1 have different effects on the development of stroma‐dependent hematopoietic cells

“…In this context, cs factors mediating cell-cell interactions may substitute for high local concentrations of a soluble factor resulting in delayed internalization of the factor/ receptor complex that normally takes place immediately after soluble factor/receptor interaction. Indeed, it was reported, that csSCF is more persistent in inducing phosphorylation of the SCF receptor than soluble SCF (Miyazawa et al, 1995). The molecular mechanisms by which the CSF-1/CSF-1 receptor complex causes different biological effects are not known.…”

Membrane‐bound Kit ligand regulates melanocyte adhesion and survival, providing physical interaction with an intraepithelial niche

“…The expression of mb-M2-KitL in Sl/Sl fibroblasts prolonged the survival of hematopoietic progenitor cells (39). This enhanced survival correlated with persistent c-kit tyrosine phosphorylation and increased cell surface expression of the c-kit receptor (40). In contrast, S-KitL induced transient phosphorylation and rapid down-regulation of c-kit, due to activity-dependent ubiquitination and degradation (41).…”

Chemokines, chemokine receptors and hematopoiesis

“…Steel factor binds the c-kit-encoded tyrosine kinase protein receptor through which it elicits its signal. While a great deal is known regarding Steel factor-mediated signaling ( (17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32)(33)(34) and references cited therein), new information continues to be published in this area. Another potent co-stimulating cytokine is Flt3-ligand (L), which signals through the Flt3 receptor and which also expresses intrinsic tyrosine kinase activity (16,(35)(36)(37)(38)(39)(40).…”