Kazerounian Shiva, Pitari Giovanni M, Shah Fawad J, Frick Glen S, Madesh Muniswamy, Ruiz-Stewart Inez, Schulz Stephanie, Hajnóczky Gyorgy, Waldman Scott A
Division of Clinical Pharmacology, Department of Medicine, Thomas Jefferson University, Philadelphia, PA 19107, USA.
J Pharmacol Exp Ther. 2005 Sep;314(3):1013-22. doi: 10.1124/jpet.105.089052. Epub 2005 Jun 3.
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.
鸟苷酸环化酶C以及细菌热稳定肠毒素（STs）诱导的cGMP积累可促进结肠癌细胞的细胞生长停滞，在肠道中起到肿瘤抑制作用。相反，通过储存性钙通道（SOCs）进行的容量性钙内流是促进结肠癌细胞增殖的关键信号机制。本研究表明，在T84人结肠癌细胞中，通过SOCs进行的容量性钙内流所介导的增殖信号与鸟苷酸环化酶C介导的细胞生长停滞相互对立且相互偶联。使用SOCs的选择性抑制剂2-氨基乙氧基二苯硼酸盐（2-APB）消除容量性钙内流，可增强ST诱导的细胞生长停滞。容量性钙内流对ST诱导的细胞生长停滞的对抗作用反映了对鸟苷酸环化酶C信号的相互抑制。钙-ATP酶抑制剂毒胡萝卜素或受体激动剂UTP或卡巴胆碱诱导的通过SOCs的钙内流抑制了鸟苷酸环化酶C依赖性cGMP的积累。钙离子载体离子霉素模拟了这种效应，而2-APB和细胞内钙螯合剂1,2-双（邻氨基-5,5'-二溴苯氧基）乙烷-N,N,N',N'-四乙酸乙酰甲酯（BAPTA-AM）可阻断这种效应。此外，容量性钙内流的调节反映了鸟苷酸环化酶C对该阳离子抑制的配体依赖性敏感化。尽管基础催化活性难以被影响，但ST刺激的鸟苷酸环化酶C受到钙的抑制，钙拮抗了镁与受体-效应器偶联所需的变构位点的结合。这些观察结果表明，通过SOCs进行的容量性钙内流对鸟苷酸环化酶C信号的相互调节代表了平衡结肠癌细胞增殖和细胞生长停滞的协调机制的一个方面。它们表明，联合使用鸟苷酸环化酶C激动剂和SOC抑制剂为结直肠肿瘤的cGMP导向治疗和预防提供了一种新的模式。
