Identification of a compound that directly stimulates phospholipase C activity
Identification of a compound that directly stimulates phospholipase C activity
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DOI：
10.1124/mol.63.5.1043
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发表时间：
2003-05-01
影响因子：
3.6
通讯作者：
Ryu, SH
中科院分区：
文献类型：
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作者：
Bae, YS;Lee, TG;Ryu, SH
磷酸肌醇特异性磷脂酶 C (PLC) 在各种细胞反应的信号转导中发挥着关键作用。然而，尽管确定 PLC 活动调节剂无疑很重要，但迄今为止尚未确定直接的 PLC 活动调节剂。在这项研究中，通过筛选人类中性粒细胞中的 10,000 多种不同化合物，我们鉴定出了一种能够强烈增强超氧化物生成活性的化合物，众所周知，这种化合物具有 PLC 依赖性。活性化合物 2,4,6-三甲基-N-(间-3-三氟甲基苯基)苯磺酰胺 (m-3M3FBS) 刺激中性粒细胞短暂的细胞内钙浓度 ([Ca2+](i)) 增加。此外，m-3M3FBS 刺激 U937 细胞中磷酸肌醇的形成，表明它刺激 PLC 活性。该化合物在包括白细胞、成纤维细胞和神经元细胞在内的各种细胞系中的 [Ca2+](i) 增加方面没有表现出细胞类型特异性。我们还通过确认以下内容排除了异三聚体 G 蛋白参与 m-3M3FBS 刺激的信号传导的可能：1) 百日咳毒素不抑制 m-3M3FBS 诱导的 [Ca2+](i) 增加； 2) m-3M3FBS不刺激环AMP的产生； 3) G 蛋白信号传导 2 调节剂对 G(q) 的抑制不影响 m-3M3FBS 诱导的 [Ca2+](i) 增加。我们还观察到 m-3M3FBS 在体外刺激 PLC 活性。所测试的纯化 PLC 亚型（即 beta2、beta3、gamma1、gamma2 和 delta1）被 m-3M3FBS 激活，并且没有表现出亚型特异性。综上所述，这些结果表明 m-3M3FBS 通过直接激活 PLC 来调节中性粒细胞功能。由于 m-3M3FBS 是已知的第一种直接激活 PLC 的化合物，因此它在研究 PLC 激活和 PLC 介导的细胞信号传导的基本分子机制中应该是有用的。
Phosphoinositide-specific phospholipase C (PLC) plays a pivotal role in the signal transduction of various cellular responses. However, although it is undeniably important that modulators of PLC activity be identified, no direct PLC activity modulator has been identified until now. In this study, by screening more than 10,000 different compounds in human neutrophils, we identified a compound that strongly enhances superoxide-generating activity, which is well known to be PLC-dependent. The active compound 2,4,6-trimethyl-N-(meta-3-trifluoromethylphenyl)benzenesulfonamide (m-3M3FBS) stimulated a transient intracellular calcium concentration ([Ca2+](i)) increase in neutrophils. Moreover, m-3M3FBS stimulated the formation of inositol phosphates in U937 cells, indicating that it stimulates PLC activity. The compound showed no cell-type specificity in terms of [Ca2+](i) increase in the various cell lines including leukocytes, fibroblasts, and neuronal cells. We also ruled out the possible involvement of heterotrimeric G proteins in m-3M3FBS-stimulated signaling by confirming the following: 1) pertussis toxin does not inhibit m-3M3FBS-induced [Ca2+](i) increase; 2) m-3M3FBS does not stimulate cyclic AMP generation; and 3) the inhibition of G(q) by the regulator of G protein-signaling 2 does not affect the m-3M3FBS-induced [Ca2+](i) increase. We also observed that m-3M3FBS stimulated PLC activity in vitro. The purified isoforms of PLC that were tested (i. e., beta2, beta3, gamma1, gamma2, and delta1) were activated by m-3M3FBS and showed no isoform specificity. Taken together, these results demonstrate that m-3M3FBS modulates neutrophil functions by directly activating PLC. Because m-3M3FBS is the first compound known to directly activate PLC, it should prove useful in the study of the basic molecular mechanisms of PLC activation and PLC-mediated cell signaling.