Gastrin Modulates Growth of a Rat Acinar Pancreatic Cell Line: Receptor Analysis and Signal Transduction

Seva, Catherine; Vries, Luc De; Scemama, Jean-Luc; Sarfati, Patrice D.; Nicolet, T; Pradayrol, Lucien; Vaysse, N.

doi:10.1159/000200381

Cited by 22 publications

(11 citation statements)

References 3 publications

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“…Further, in heterodimer-expressing cells, agonist-induced calcium responses were poorly inhibited by both antagonists. Indeed, type A and type B CCK receptors have been found to co-localize in select areas of the brain where opioids have effects (30), and both of these receptors have been described to be expressed on AR4 -2J cells (31). Our data suggest that type A and type B CCK heterodimers could provide an explanation for the above observations.…”

Section: Discussionsupporting

confidence: 60%

Heterodimerization of Type A and B Cholecystokinin Receptors Enhance Signaling and Promote Cell Growth

Cheng

Harikumar

Holicky

et al. 2003

Journal of Biological Chemistry

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G protein-coupled receptors represent the largest and most diverse superfamily of transmembrane receptors and mediate the effects of a wide variety of extracellular stimuli. It is now well established that, like single transmembrane tyrosine kinase receptors (2), these heptahelical receptors can form oligomeric complexes in the plasma membrane (3-13). Evidence for this includes biochemical, biophysical, and functional data (3, 6, 7). In addition to homodimerization, heterodimerization can occur with some structurally related receptors in this superfamily (8 -10, 12). This has been implicated in the effects on ligand binding and acute effects on cellular signaling (8,9,12). Formation of heterodimers of nonfunctional B1 and B2 ␥-aminobutyric acid receptors have been shown to be necessary for binding and complete functional activity of native ligands (10). Dimeric complexes of fully functional opioid receptors have been shown to alter their selectivity, displaying decreased affinities for highly selective agonists and enhanced affinities for nonselective agonists (9). Dimerization between somatostatin receptors has also been found to alter the pharmacology and signaling of individual receptors (12). However, little is known about the relevance of receptor dimerization to longer term effects, such as cell growth.Gastrin and cholecystokinin (CCK) 1 are gastrointestinal and neuronal peptides with important regulatory roles in the digestive tract and nervous system, including both acute and more chronic trophic effects (14 -16). These functions are mediated by two receptors, the type A and type B CCK receptors, both belonging to the class I family of G protein-coupled receptors and exhibiting 48% structural homology with each other (17, 18). These two receptors can be distinguished on the basis of their structural specificity, with both recognizing gastrin and CCK but with each having markedly distinct sensitivities to the state of tyrosine sulfation of those peptides.Although additional subtypes of CCK receptors have been predicted to exist, based on the complexity of the pharmacology and physiology of gastrin and CCK in vivo, only these two CCK receptor cDNAs have thus far been identified. It is possible that receptor oligomerization contributes to the pharmacology that has been observed and may even play a role in cell growth. We recently utilized bioluminescence resonance energy transfer (BRET) and immunoprecipitation to demonstrate that type A CCK receptors can exist as homodimers in living cells, with these complexes dissociated by agonist binding (1). To date there have been no reports of the ability of type B CCK receptors to dimerize or the possibility of the type A and B CCK receptors to form heterodimeric complexes.Indeed, using the same biophysical techniques, our current study demonstrated homodimerization of type B CCK receptors, as well as heterodimerization of this receptor with type A CCK receptors when co-expressed in cells. Of note, unlike the type A receptors (1), oligomerization of the type B CCK r...

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Section: Discussionsupporting

confidence: 60%

Heterodimerization of Type A and B Cholecystokinin Receptors Enhance Signaling and Promote Cell Growth

Cheng

Harikumar

Holicky

et al. 2003

Journal of Biological Chemistry

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“…Maximal binding sites (B max ) of respectively 29+6 fmoles (clone 1), 18.4+2 fmoles (clone 15) and 28+5 fmoles (clone 19) per 10 6 cells, demonstrated an expression level comparable to that of the pancreatic tumor cell line, AR4-2J, that endogenously expresses the CCK-2R. The cells possessed CCK-specific highaffinity binding sites at their surface (Kd 0.56+0.1 nM (clone 1), 0.51+0.06 nM (clone 15), 0.65+0.1 nM (clone 19)), again similar to those of endogenously expressed CCK-2R in AR4-2J cells (Seva et al, 1990;C Seva, unpublished). No specific binding was detected on parental cells.…”

Section: Resultsmentioning

confidence: 67%

“…Binding experiments were performed as described previously (Seva et al, 1990). Briefly, 1.25610 5 cells seeded into 24-well plates were incubated with 50 pM of 125 I-BH-(Thr, Nle-CCK1-9s) with or without excess of unlabeled gastrin.…”

Section: Binding Experimentsmentioning

confidence: 99%

Gastrin mediated cholecystokinin-2 receptor activation induces loss of cell adhesion and scattering in epithelial MDCK cells

et al. 2002

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The presence of gastrin and CCK-2/gastrin receptors in human preneoplastic and neoplastic lesions of pancreas and colon suggests a role in cancer development. Gastrin's growth-promoting action has been established, but a role in cellular morphogenetic processes promoting tumor invasion has been elusive. Our aim was (i) to investigate whether activation of the CCK-2R affects cellular morphology, intercellular adhesion and motility, as crucial parameters of epithelial differentiation, and (ii) to identify the signaling pathways and mechanisms implicated. Madin-Darby Canine Kidney (MDCK) cells were chosen to generate an epithelial non-tumorigenic model system expressing human CCK-2R. Epithelial differentiation and motility were analysed upon CCK-2R activation using immunocytochemistry and invasion assays. The functionality of adhesion complexes and activity of signaling proteins was determined with biochemical techniques. CCK-2R activation induced cell dissociation and enhanced invasion, preceded by decreased membrane localization of adherens junction molecules and nuclear accumulation of b-catenin. Concomitantly, and requiring the activation of several signaling pathways, catenins were shifted from the cytoskeletal to the cytoplasmic fraction, suggesting the detachment of the cytoskeleton from the adherens complex. These data represent the first evidence for the CCK-2R, regulating cell -cell and cell -substrate adhesion and support a role for CCK-2R in the progression of carcinoma.

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“…Specific culture conditions involving synchronisation of cells with thymidine and growth in serum-free medium have been reported previously to be important in revealing trophic effects of gastrin on gastric and colon cancer cells in vitro (Kusyk et al, 1986;Watson et al, 1988;Guo et al, 1990). In previous studies, ornithine decarboxylase activity, or 3H-thymidine or 75Se-selenomethionine incorporation were used as indices of responsiveness of AR4-2J cells to gastrin (Scemama et al, 1989;Seva et al, 1990;Watson et al, 1991a). Interpretation of data obtained by the latter approach is frustrated by the fact that gastrin increases secretion, and presumably synthesis, of proteins such as the enzyme amylase from AR4-2J cells (Lambert et al, 1991).…”

Section: Discussionmentioning

confidence: 99%

Autocrine stimulation of growth of AR4-2J rat pancreatic tumour cells by gastrin

Blackmore

Bh²

1992

Br J Cancer

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Summary The control of cell proliferation by gastrin has been investigated in a rat pancreatic tumour cell line, AR4-2J. Exogenous gastrin, 10-12 to 10-8 M, stimulated cell growth of thymidine-synchronised AR4-2J cells cultured over 48 h in serum-free medium. Cell lysates of AR4-2J cells contained an average of 4.5 and 3.5 pg gastrin per 106 cells, when grown in serum-supplemented or serum-free media, respectively, as revealed by radioimmunoassay. In serum-free medium, AR4-2J secrete 34ngl 10-6 cells of gastrin over 48h. Addition of an anti-gastrin immunoglobulin preparation, but not control immunoglobulins, caused a maximum 52% reduction in cell growth. These data are consistent with an autocrine role for gastrin in the control of AR4-2J cell growth. These results were supported by studies with gastrin/CCK receptor antagonists. Six non-peptide gastrin/CCK receptor antagonists inhibited AR4-2J cell growth in a concentration-related manner. The concentration required for 50% inhibition (IC50) of cell growth by the amino acid-derived antagonists proglumide (3.5 x 10-M), benzotript (1.8 x 10-3 M), loxiglumide (1.1 x 10-4 M) and lorglumide (6.7 x 10-5 M) were of the same order and significantly correlated with their IC50 for inhibition of 1251-gastrin binding to AR4-2J cells. Inhibition of cell growth by these antagonists was partially reversed by the addition of exogenous gastrin. In contrast, the ICm for inhibition of cell growth with two benzodiazepine-derived antagonists, the CCK-B receptor antagonist L-365,260 (4.6 x 10-5 M) and the CCK-A receptor antagonist devazepide (1.7 x 10-' M) were two-three orders of magnitude greater than those required to inhibit gastrin binding (10-8-10-7 M). The growth inhibitory effects of L-365,260 and devazepide were not reversed by exogenous gastrin suggesting these benzodiazepine-derived antagonists do not inhibit cell growth by interaction with gastrin receptors. The results are consistent with gastrin being an autocrine growth factor in AR4-2J cells, and that stimulation of cell growth is due to stimulation of the gastrin, rather than CCK-B, receptor sub-type. This study highlights that gastrin receptor antagonists warrant further investigation as agents to control growth of tumours, such as those from the gastrointestinal tract, which express gastrin receptors.

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Gastrin Modulates Growth of a Rat Acinar Pancreatic Cell Line: Receptor Analysis and Signal Transduction

Cited by 22 publications

References 3 publications

Heterodimerization of Type A and B Cholecystokinin Receptors Enhance Signaling and Promote Cell Growth

Heterodimerization of Type A and B Cholecystokinin Receptors Enhance Signaling and Promote Cell Growth

Gastrin mediated cholecystokinin-2 receptor activation induces loss of cell adhesion and scattering in epithelial MDCK cells

Autocrine stimulation of growth of AR4-2J rat pancreatic tumour cells by gastrin

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