Claudin-2 suppresses GEF-H1, RHOA, and MRTF, thereby impacting proliferation and profibrotic phenotype of tubular cells
Claudin-2 suppresses GEF-H1, RHOA, and MRTF, thereby impacting proliferation and profibrotic phenotype of tubular cells
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DOI：
10.1074/jbc.ra118.006484
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发表时间：
2019-10-18
影响因子：
4.8
通讯作者：
Szaszi, Katalin
中科院分区：
文献类型：
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作者：
Dan, Qinghong;Shi, Yixuan;Szaszi, Katalin
紧密连接的致孔蛋白Claudin-2(CLDN-2)介导渗漏上皮细胞旁Na+和水的转运，改变癌细胞的增殖。此前，我们报道了肿瘤坏死因子-？时间依赖性地改变肾小管上皮细胞CLDN-2的表达。在这里，我们在小鼠肾损伤模型(单侧输尿管梗阻)中发现了类似的表达模式，包括最初的增加和随后的CLDN-2蛋白表达的下降。在LLC-PK1肾小管上皮细胞中，CldN-2沉默可诱导鸟嘌呤核苷酸交换因子H1的激活和磷酸化，导致RAS同源家族成员A(RHOA)的激活。沉默其他Claudins则没有这种作用，而抗siRNA的CLDN-2的重新表达阻止了RHOA的激活，表明CLDN-2对RHOA具有特异性作用。此外，CLDN-2基因敲除小鼠的肾脏中活性RHOA水平升高。值得注意的是，CLDN-2沉默降低了LLC-PK1细胞的增殖，并上调了细胞周期蛋白依赖性激酶抑制物P27(P27KIP1)的表达，这种抑制作用依赖于环境基金-H1/RHOA。P27KIP1沉默消除了CLDN-2缺失对细胞增殖的影响。CLDN-2缺失还激活了肌钙蛋白相关转录因子(MRTF)，这是一种致纤维化的RHOA效应因子，并增加了结缔组织生长因子和平滑肌肌动蛋白的表达。CLDN-2下调参与了肿瘤坏死因子-1诱导的RHOA活化和平滑肌肌动蛋白表达。治疗，因为CLDN-2的重新表达减轻了它们的影响。我们的结果表明，CLDN-2抑制了全球环境基金-H1/RHOA。例如，通过炎症下调CLDN-2可以减少增殖，并通过RHOA促进MRTF的激活。这些发现表明，最初CLDN-2的升高可能有助于上皮再生，而CLDN-2的缺失可能有助于纤维化的重新编程。
The tight junctional pore-forming protein claudin-2 (CLDN-2) mediates paracellular Na+ and water transport in leaky epithelia and alters cancer cell proliferation. Previously, we reported that tumor necrosis factor-? time-dependently alters CLDN-2 expression in tubular epithelial cells. Here, we found a similar expression pattern in a mouse kidney injury model (unilateral ureteral obstruction), consisting of an initial increase followed by a drop in CLDN-2 protein expression. CLDN-2 silencing in LLC-PK1 tubular cells induced activation and phosphorylation of guanine nucleotide exchange factor H1 (GEF-H1), leading to Ras homolog family member A (RHOA) activation. Silencing of other claudins had no such effects, and re-expression of an siRNA-resistant CLDN-2 prevented RHOA activation, indicating specific effects of CLDN-2 on RHOA. Moreover, kidneys from CLDN-2 knockout mice had elevated levels of active RHOA. Of note, CLDN-2 silencing reduced LLC-PK1 cell proliferation and elevated expression of cyclin-dependent kinase inhibitor P27 (P27KIP1) in a GEF-H1/RHOA-dependent manner. P27KIP1 silencing abrogated the effects of CLDN-2 depletion on proliferation. CLDN-2 loss also activated myocardin-related transcription factor (MRTF), a fibrogenic RHOA effector, and elevated expression of connective tissue growth factor and smooth muscle actin. Finally, CLDN-2 down-regulation contributed to RHOA activation and smooth muscle actin expression induced by prolonged tumor necrosis factor-? treatment, because they were mitigated by re-expression of CLDN-2. Our results indicate that CLDN-2 suppresses GEF-H1/RHOA. CLDN-2 down-regulation, for example, by inflammation, can reduce proliferation and promote MRTF activation through RHOA. These findings suggest that the initial CLDN-2 elevation might aid epithelial regeneration, and CLDN-2 loss could contribute to fibrotic reprogramming.