CD9 regulates keratinocyte migration by negatively modulating the sheddase activity of ADAM17
CD9 regulates keratinocyte migration by negatively modulating the sheddase activity of ADAM17
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CD9 通过负调节 ADAM17 的脱落酶活性来调节角质形成细胞迁移
DOI：
10.7150/ijbs.29404
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发表时间：
2019-01-01
影响因子：
9.2
通讯作者：
Zhang, Jiaping
中科院分区：
文献类型：
--
作者：
Liu, Jie;Zhu, Guoqin;Zhang, Jiaping
CD9是一种跨膜蛋白，最近参与了不同的生理和细胞过程，如细胞迁移和黏附。根据先前的研究，CD9的下调有助于角质形成细胞的迁移，这对伤口的再上皮化至关重要。然而，人们普遍认为Tetraspanin CD9没有配体或作为细胞表面受体的功能，而是被认为与其他跨膜分子结合，从而介导角质形成细胞的迁移。CD9是如何与迁移角质形成细胞中的跨膜分子相关联的，目前知之甚少。在此，我们利用共聚焦显微镜观察到，在体内和体外的创伤修复过程中，Tetraspanin CD9和ADAM17共定位于角质形成细胞的表面。免疫共沉淀实验表明，CD9和ADAM17在HaCaT细胞和C57-MKs中存在直接关联。功能研究表明，CD9的下调或过度表达对ADAM17脱落酶活性具有负性调节作用。这种活性参与了CD9调节的细胞运动和迁移。进一步研究发现，ADAM17抑制剂TAPI-2或siADAM17可显著阻断CD9下调诱导的角质形成细胞迁移的增强效应。同时，TAPI-2抑制ADAM17的脱落酶活性，从而减少CD9沉默的HaCat细胞和C57-MKs中AREG和HB-EGF的释放。重要的是，抗HB-EGF的中和抗体显著降低CD9沉默的角质形成细胞的迁移和运动能力，而siADAM17对CD9调节的角质形成细胞迁移的抑制可通过加入重组HB-EGF而解除，激活EGFR/ERK通路。综上所述，我们的结果表明，ADAM17脱落酶活性是通过下调CD9而激活的，从而介导HB-EGF的脱落和EGFR/ERK信号的激活，这对角质形成细胞的迁移和伤口愈合具有至关重要的影响。
CD9 is a trans-membrane protein, and has recently been implicated in different physiological and cellular processes, such as cell migration and adhesion. According to previous study, down-regulation of CD9 contributes to keratinocyte migration, critical for wound re-epithelialization. Nevertheless, it is widely believed that tetraspanin CD9 does not have ligands or function as the cell surface receptor, rather it is thought to associate with other transmembrane molecules, thereby mediate keratinocyte migration. Little is known about how CD9 associates with transmembrane molecules in migratory keratinocytes. Here, using confocal microscopy, we observed that tetraspanin CD9 and ADAM17 co-localized on the surface of keratinocytes in the course of wound repair in vivo and in vitro. Co-immunoprecipitation experiments demonstrated a direct association between CD9 and ADAM17 in HaCaT cells and C57-MKs. Functional studies revealed that down-regulation or over-expression of CD9 exerted negative regulatory effects on ADAM17 sheddase activity. This activity is involved in CD9-regulated cell motility and migration. Further studies found that ADAM17 inhibitor-TAPI-2 or siADAM17 significantly abolished the enhanced effect of keratinocyte migration induced by CD9 down-regulation. Meanwhile, the sheddase activity of ADAM17 was inhibited by TAPI-2, which decreased this release of AREG and HB-EGF in CD9-silenced HaCat cells and C57-MKs. Importantly, neutralizing antibody against HB-EGF significant weakened keratinocyte migration and motility in CD9-silenced keratinocytes, and the inhibition of CD9-regulated keratinocyte migration by siADAM17 was rescued by addition of recombinant HB-EGF, activating EGFR/ERK pathway. Collectively, our results suggest that ADAM17 sheddase activity is activated by down-regulation of CD9, thereby mediating shedding of HB-EGF and activation of EGFR/ERK signaling, which crucially affects the keratinocyte migration and wound healing.