Abstract
Intracellular mechanisms that mediate mitogenic effects of gastrin remain largely unknown. The present studies were designed to examine if protein tyrosine kinase (PTKs) mediate growth effects of gastrin on a rat intestinal epithelial cell line (IEC-6 cells). Gastrin (<10 nM) was mitogenic for IEC-6 cells. PTK activity of cell membranes was stimulated in response to 0.01- 10.0 nM and 0.05-10.0 μM gastrin in a double biphasic manner. Cells labeled with H332PO4 were stimulated with gastrin and cellular proteins immunoprecipitated with phosphotyrosine antibodies. Endogenous proteins were phosphorylated in a dose- (100% effective dose = 0.1-1.0 nM) and time- dependent manner; at >10 nM gastrin, the second peak of response was not measured in intact cells. Thus the growth and phosphorylation response of intact cells to gastrin was similar. Both high [dissociation constant (K(d)) = 1 nM] and low (K(d) = ~0.1 μM)-affinity gastrin binding sites are present on IEC-6 cells. The results of the present study suggest, that occupancy of both high- and low-affinity gastrin-binding sites can potentially activate membrane-associated PTKs. However, in intact cells, occupancy of low-affinity sites apparently attenuates kinase activity resulting in reduced protein phosphorylation. Eight protein bands [with relative molecular weight (M(r)) of 32-145 kDa] were tyrosine phosphorylated in intact cells in response to 0.1-1.0 nM gastrin, including two pp60 src-like proteins (with M(r) of 54 and 62 kDa). Thus the growth response pattern of a target cell to gastrin may depend on the stimulation of kinases and other factors (phosphatases?) that phosphorylate and/or dephosphorylate several proteins including c-src-like proteins in a dose-dependent manner.
Original language | English (US) |
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Pages (from-to) | G235-G244 |
Journal | American Journal of Physiology - Gastrointestinal and Liver Physiology |
Volume | 267 |
Issue number | 2 30-2 |
DOIs | |
State | Published - 1994 |
Externally published | Yes |
Keywords
- IEC-6 cells
- gastrin receptor
- in vitro
- mitogenic effects
- tyrosine kinases
ASJC Scopus subject areas
- Physiology
- Hepatology
- Gastroenterology
- Physiology (medical)