The effects of s analgus RC-160and SMS-201-995 on tyrosine phosphatase and cell proliferation were investigated in COS-7 and NIH 3T3 cells expressing human somatostatin receptor subtype 1 or 2 (SSTR1 or SSTR2 antagonize the mitogenic effect of growth factors acting on tyrosine kinase receptors such as epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) (9-11). Furthermore, these analogues have been found to stimulate tyrosine phosphatase activity in normal and tumoral pancreatic cells (9,(12)(13)(14)(15) and to activate the dephosphorylation of EGF receptor (9, 16). The ability of somatostatin analogues to stimulate tyrosine phosphatase correlates with their inhibitory effect on pancreatic cell growth, and this correlation supports the hypothesis that the growth inhibition is mediated by dephosphorylation of tyrosine protein signals. Somatostatin analogues might suppress tumor growth by reversing the stimulatory effect of EGF on phosphorylation of EGF receptor tyrosine kinase and EGF-phosphorylated proteins (17). We also observed that a membrane tyrosine phosphatase is coeluted with somatostatin receptor, suggesting that tyrosine phosphatase may be a part ofthe signal transduction pathway promoted by somatostatin receptor occupancy (16).The somatostatin receptor subtypes and the molecular mechanism involved in the tyrosine phosphatase stimulation have been, until now, unknown (18, 19). We must better understand what physiological response every subtype elicits, how their signals are processed in the cell, and in what normal and/or pathological tissues each is expressed to choose the appropriate analogue for targeting to specific cells for therapeutic use.In the present study, we examined the effects of two somatostatin analogues, RC-160 and SMS, on binding and signal transduction pathways of the two human cloned somatostatin receptor subtypes hSSTR1 and hSSTR2 transiently expressed in COS-7 cells. We also investigated the 2315The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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Activation of the somatostatin receptor sst2, a member of the G i protein-coupled receptor family, results in the stimulation of a protein-tyrosine phosphatase activity involved in the sst2-mediated growth inhibitory signal. Here, we report that SHP-1, a cytoplasmic proteintyrosine phosphatase containing two Src homology 2 domains constitutively associated with sst2 as evidence by coprecipitation of SHP-1 protein with sst2, in Chinese hamster ovary cells coexpressing sst2 and SHP-1. Activation of sst2 by somatostatin resulted in a rapid dissociation of SHP-1 from sst2 accompanied by an increase of SHP-1 activity. SHP-1 was phosphorylated on tyrosine in control cells and somatostatin induced a rapid and transient dephosphorylation on tyrosine residues of the enzyme. Stimulation of SHP-1 activity by somatostatin was abolished by pertussis toxin pretreatment of cells. G i␣3 was specifically immunoprecipitated by anti-sst2 and anti-SHP-1 antibodies, and somatostatin induced a rapid dissociation of G i␣3 from sst2, suggesting that G i␣3 may be involved in the sst2⅐SHP-1 complexes. Finally, somatostatin inhibited the proliferation of cells coexpressing sst2 and SHP-1, and this effect was suppressed in cells coexpressing sst2 and the catalytic inactive SHP-1 (C453S mutant). Our data identify SHP-1 as the tyrosine phosphatase associated with sst2 and demonstrate that this enzyme may be an initial key transducer of the antimitogenic signaling mediated by sst2.

We investigated cell proliferation modulated by cholecystokinin (CCK) and somatostatin analogue RC-160 in CHO cells bearing endogenous CCK A receptors and stably transfected by human subtype sst5 somatostatin receptor. CCK stimulated cell proliferation of CHO cells. This effect was suppressed by inhibitor of the soluble guanylate cyclase, LY 83583, the inhibitor of the cGMP dependent kinases, KT 5823, and the inhibitor of mitogen-activated protein (MAP) kinase kinase, PD 98059. CCK treatment induced an increase of intracellular cGMP concentrations, but concomitant addition of LY 83583 virtually suppressed this increase. CCK also activated both phosphorylation and activity of p42-MAP kinase; these effects were inhibited by KT 5823. All the effects of CCK depended on a pertussis toxin-dependent G protein. Somatostatin analogue RC-160 inhibited CCK-induced stimulation of cell proliferation but it did not potentiate the suppressive effect of the inhibitors LY 83583 and KT 5823. RC-160 inhibited both CCK-induced intracellular cGMP formation as well as activation of p42-MAP kinase phosphorylation and activity. This inhibitory effect was observed at doses of RC-160 similar to those necessary to occupy the sst5 recombinant receptor and to inhibit CCK-induced cell proliferation. We conclude that, in CHO cells, the proliferation and the MAP kinase signaling cascade depend on a cGMPdependent pathway. These effects are positively regulated by CCK and negatively inf luenced by RC-160, interacting through CCK A and sst5 receptors, respectively. These studies provide a characterization of the antiproliferative signal mediated by sst5 receptor.Somatostatin and cholecystokinin are both regulatory hormones widely distributed in the body. Both these hormones participate in a variety of biological processes including neurotransmission and control of pancreatic secretion. They induce their biological effects by interacting with specific G protein-linked receptors (1, 2). The role of the two peptides in the regulation of cell growth also has been demonstrated in studies carried out both in vitro and in vivo (2-6).The effects of CCK and its related peptide, gastrin, are transduced through at least two distinct subtypes of specific cell-surface receptors, CCK A (alimentary) and CCK B (brain, also called CCK B ͞gastrin receptors), that have been cloned in human and rodents (1). CCK B ͞gastrin receptors bind CCK and gastrin with the same high affinity. In contrast, the CCK A receptors exhibit a 100-to 500-fold higher affinity for CCK than gastrin. Gastrin has trophic effects on normal digestive mucosa and stimulates the growth of colon, gastric, and pancreatic cancer cell lines in culture (7,8). This proliferative effect has been shown to be mediated by the gastrin͞CCK B receptors in different cell models (8, 9).In the rat exocrine pancreas that expresses CCK A type receptor, CCK is known to stimulate pancreatic growth both in vitro and in vivo (3-5). The CCK A receptor interacts with G proteins and thereby activates phospholipas...