Singh Bhuminder, Schneider Matthias, Knyazev Pjotr, Ullrich Axel
Department of Molecular Biology, Max-Planck-Institute of Biochemistry, Martinsried, Germany.
Int J Cancer. 2009 Feb 1;124(3):531-9. doi: 10.1002/ijc.23974.
Exposure to extensive ultraviolet (UV) rays is a major cause of skin cancer, which is thought to be initiated by DNA mutations. Members of the epidermal growth factor receptor (EGFR) family are important in various pathophysiologic processes like cancer and are shown to be phosphorylated upon UV exposure. Here we show that EGFR phosphorylation by modest UV doses is dependent on metalloprotease activity and resultant epidermal growth factor (EGF) family proligand shedding. This proligand cleavage releases the mature ligand, which then binds to and activates EGFR. We show that UV induced EGFR phosphorylation in transformed cell lines of melanocyte and keratinocyte origin, which was reduced upon preincubation with a broad-spectrum metalloprotease inhibitor, BB94. UV also activated EGFR downstream signaling via Erk and Akt pathways in a BB94-sensitive manner. Furthermore, using neutralizing antibodies we found that proligand amphiregulin was required for UV-induced EGFR activation in SCC-9 cells. Using RNAi this EGFR activation was further shown to depend on the metalloproteases ADAM9 and ADAM17 in SCC-9 cells. cDNA array hybridization and RT-PCR analysis showed overexpression of a Disintegrin and a Metalloproteases (ADAMs) and EGF family proligands in melanoma cell lines. Additionally, blocking EGFR signal transactivation by BB94 led to increased apoptosis in UV-irradiated cells. EGFR signal transactivation also led to increased stability of the DNA repair protein, PARP, under UV stress. Thus, both antiapoptotic and DNA repair pathways are activated simultaneously by EGFR signal transactivation. Together, our data provide novel insights into the mechanism of UV-induced EGFR activation, suggesting broad relevance of the UV-ADAM-proligand-EGFR-Erk/Akt pathway and its significance in skin cancer.
暴露于大量紫外线（UV）是皮肤癌的主要病因，皮肤癌被认为是由DNA突变引发的。表皮生长因子受体（EGFR）家族成员在癌症等多种病理生理过程中起重要作用，且已表明在紫外线暴露后会发生磷酸化。在此我们表明，适度紫外线剂量引起的EGFR磷酸化依赖于金属蛋白酶活性以及由此导致的表皮生长因子（EGF）家族前体配体的脱落。这种前体配体的切割释放出成熟配体，然后该配体结合并激活EGFR。我们发现，紫外线在源自黑素细胞和角质形成细胞的转化细胞系中诱导EGFR磷酸化，而在与广谱金属蛋白酶抑制剂BB94预孵育后这种磷酸化会减少。紫外线还以BB94敏感的方式通过Erk和Akt途径激活EGFR下游信号传导。此外，使用中和抗体我们发现，前体配体双调蛋白是SCC - 9细胞中紫外线诱导的EGFR激活所必需的。使用RNA干扰进一步表明，SCC - 9细胞中的这种EGFR激活依赖于金属蛋白酶ADAM9和ADAM17。cDNA阵列杂交和RT - PCR分析表明，在黑色素瘤细胞系中，解整合素和金属蛋白酶（ADAMs）以及EGF家族前体配体过表达。此外，BB94阻断EGFR信号反式激活导致紫外线照射细胞的凋亡增加。在紫外线应激下，EGFR信号反式激活还导致DNA修复蛋白PARP的稳定性增加。因此，抗凋亡和DNA修复途径通过EGFR信号反式激活同时被激活。总之，我们的数据为紫外线诱导EGFR激活的机制提供了新的见解，表明紫外线 - ADAM - 前体配体 - EGFR - Erk/Akt途径具有广泛相关性及其在皮肤癌中的重要性。
