Department of Biomedical Science, Jungwon University, Goesan, Chungcheongbukdo 367-805, Korea.
J Microbiol Biotechnol. 2012 Feb;22(2):170-5. doi: 10.4014/jmb.1110.10075.
Clostridium difficile toxin A glucosylates Rho family proteins, resulting in actin filament disaggregation and cell rounding in cultured colonocytes. Given that the cellular toxicity of toxin A is dependent on its receptor binding and subsequent entry into the cell, we herein sought to identify additional colonocyte proteins that might bind to toxin A following its internalization. Our results revealed that toxin A interacted with ERK1 and ERK2 in two human colonocyte cell lines (NCM460 and HT29). A GST-pulldown assay also showed that toxin A can directly bind to ERK1 and ERK2. In NCM460 cells exposed to PMA (an ERK1/2 activator), the phosphorylation of ERK1/2 did not affect the interaction between toxin A and ERK1/2. However, an in vitro kinase assay showed that the direct binding of toxin A to ERK1 or ERK2 inhibited their kinase activities. These results suggest a new molecular mechanism for the cellular toxicity seen in cells exposed to toxin A.
艰难梭菌毒素 A 使 Rho 家族蛋白发生葡萄糖基化，导致培养的结肠细胞中的肌动蛋白丝解聚和细胞圆化。鉴于毒素 A 的细胞毒性依赖于其受体结合和随后进入细胞，我们在此试图鉴定可能在其内化后与毒素 A 结合的其他结肠细胞蛋白。我们的结果表明，毒素 A 在两种人结肠细胞系（NCM460 和 HT29）中与 ERK1 和 ERK2 相互作用。GST 下拉测定还表明，毒素 A 可以直接与 ERK1 和 ERK2 结合。在暴露于 PMA（ERK1/2 激活剂）的 NCM460 细胞中，ERK1/2 的磷酸化并不影响毒素 A 与 ERK1/2 之间的相互作用。然而，体外激酶测定表明毒素 A 与 ERK1 或 ERK2 的直接结合抑制了它们的激酶活性。这些结果为暴露于毒素 A 的细胞中观察到的细胞毒性提供了一个新的分子机制。
