Abstract
Guanylyl cyclase C and accumulation of cGMP induced by bacterial heat-stable enterotoxins (STs) promote colon cancer cell cytostasis, serving as a tumor suppressor in intestine. Conversely, capacitative calcium entry through store-operated calcium channels (SOCs) is a key signaling mechanism that promotes colon cancer cell proliferation. The present study revealed that proliferative signaling by capacitative calcium entry through SOCs opposes and is reciprocally coupled to cytostasis mediated by guanylyl cyclase C in T84 human colon carcinoma cells. Elimination of capacitative calcium entry employing 2-aminoethoxydiphenylborate (2-APB), a selective inhibitor of SOCs, potentiated cytostasis induced by ST. Opposition of ST-induced cytostasis by capacitative calcium entry reflects reciprocal inhibition of guanylyl cyclase C signaling. Calcium entry through SOCs induced by the calcium-ATPase inhibitor thapsigargin or the receptor agonists UTP or carbachol inhibited guanylyl cyclase C-dependent cGMP accumulation. This effect was mimicked by the calcium ionophore ionomycin and blocked by 2-APB and intracellular 1,2-bis(o-amino-5,5′-dibromophenoxy)ethane-N,N,N′,N′-tetraacetic acid-acetoxymethyl ester (BAPTA-AM), a chelator of calcium. Moreover, regulation by capacitative calcium entry reflected ligand-dependent sensitization of guanylyl cyclase C to inhibition by that cation. Although basal catalytic activity was refractory, ST-stimulated guanylyl cyclase C was inhibited by calcium, which antagonized binding of magnesium to allosteric sites required for receptor-effector coupling. These observations demonstrate that reciprocal regulation of guanylyl cyclase C signaling by capacitative calcium entry through SOCs represents one limb of a coordinated mechanism balancing colon cancer cell proliferation and cytostasis. They suggest that combining guanylyl cyclase C agonists and SOC inhibitors offers a novel paradigm for cGMP-directed therapy and prevention for colorectal tumors.
Footnotes
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This study was supported by National Institutes of Health Grants HL59214, CA75123, CA79663, and CA95026 (to S.A.W.); GM59419 (to G.H.); the Landenberger Foundation (to G.M.P.); and Targeted Diagnostics and Therapeutics, Inc. I.R.-S. was supported by National Institutes of Health Minority Supplement HL59214-0151. G.S.F. was enrolled in the National Institutes of Health Institutional K30 Training Program in Human Investigation (K30 HL004522) and was supported by National Institutes of Health Institutional Award T32 GM08562 for Postdoctoral Training in Clinical Pharmacology. S.A.W. is the Samuel M. V. Hamilton Professor of Medicine of Thomas Jefferson University.
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doi:10.1124/jpet.105.089052.
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ABBREVIATIONS: ETEC, enterotoxigenic E. coli; ST, heat-stable enterotoxin; CNG, cyclic nucleotide-gated; SOC, store-operated calcium channel; BAPTA, 1,2-bis(o-amino-5,5′-dibromophenoxy)ethane-N,N,N′,N′-tetraacetic acid; AM, acetoxymethyl ester; 2-APB, 2-aminoethoxydiphenylborate; IBMX, isobutylmethylxanthine; sGC, soluble guanylyl cyclase.
- Received May 3, 2005.
- Accepted June 1, 2005.
- The American Society for Pharmacology and Experimental Therapeutics
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