INSERM U773, Faculté de Médecine Xavier Bichat 75018 Paris, France.
Am J Pathol. 2010 Mar;176(3):1452-61. doi: 10.2353/ajpath.2010.090523. Epub 2010 Jan 7.
Certain serine proteases are considered to be signaling molecules that act through protease-activated receptors (PARs). Our recent studies have implicated PAR1 and PAR4 (thrombin receptors) and PAR2 (trypsin receptor) in human colon cancer growth. Here we analyzed the expression of KLK4, a member of the kallikrein-related peptidase (KLK) family of serine proteases and explored whether this member can activate PAR1 and PAR2 in human colon cancer cells. Immunohistochemistry showed KLK4 expression in human colon adenocarcinomas and its absence in normal epithelia. KLK4 (1 micromol/L) initiated loss of PAR1 and PAR2 from the HT29 cell surface as well as increased intracellular calcium transients in HT29 cells. This KLK4-induced Ca2+ flux was abrogated after an initial challenge of the cells with TRAP (SFLLR-NH2; 100 micromol/L), which is known to desensitize PAR1 and PAR2. Interestingly, PAR1 blocking antibody, which inhibits cleavage and activation by thrombin, dramatically reduced KLK4-induced Ca2+ influx, but blocking cleavage of PAR2 failed to attenuate the KLK4-induced Ca2+ flux. Consistently, desensitization with AP1 (TFFLR-NH2), targeting PAR1, attenuated most of the Ca2+ flux induced by KLK4. KLK4 also induced a rapid and significant ERK1/2 phosphorylation in HT29 cells. Our results demonstrate, for the first time, that KLK4 is aberrantly expressed in colon cancer and capable of inducing PAR1 signaling in cancer cells. These data suggest that KLK4 signaling via PAR1 may represent a novel pathway in colon tumorigenesis.
某些丝氨酸蛋白酶被认为是通过蛋白酶激活受体（PAR）发挥作用的信号分子。我们最近的研究表明，PAR1 和 PAR4（凝血酶受体）和 PAR2（胰蛋白酶受体）参与了人结肠癌的生长。在这里，我们分析了激肽释放酶相关肽酶（KLK）家族丝氨酸蛋白酶成员 KLK4 的表达，并探讨了该成员是否可以激活人结肠癌细胞中的 PAR1 和 PAR2。免疫组织化学显示 KLK4 在人结肠腺癌中的表达及其在正常上皮组织中的缺失。KLK4（1µmol/L）引发 HT29 细胞表面 PAR1 和 PAR2 的丢失以及 HT29 细胞内钙离子的瞬变增加。这种 KLK4 诱导的 Ca2+ 流在细胞用 TRAP（SFLLR-NH2；100µmol/L）进行初始挑战后被阻断，TRAP 已知可使 PAR1 和 PAR2 脱敏。有趣的是，PAR1 阻断抗体可抑制凝血酶的裂解和激活，可显著减少 KLK4 诱导的 Ca2+ 内流，但阻断 PAR2 的裂解并不能减弱 KLK4 诱导的 Ca2+ 流。一致地，针对 PAR1 的 AP1（TFFLR-NH2）脱敏减弱了 KLK4 诱导的大部分 Ca2+ 流。KLK4 还在 HT29 细胞中迅速诱导 ERK1/2 的磷酸化。我们的研究结果首次表明，KLK4 在结肠癌中异常表达，并且能够在癌细胞中诱导 PAR1 信号。这些数据表明 KLK4 通过 PAR1 信号可能代表结肠癌发生的一种新途径。
