Lin Fu, Chengyao Xie, Qingchang Li, Qianze Dong, Enhua Wang, Yan Wang
Department of Pathology, First Affiliated Hospital and College of Basic Medical Sciences, China Medical University, Shenyang, Liaoning, China.
Mol Carcinog. 2015 Jun;54 Suppl 1:E35-44. doi: 10.1002/mc.22148. Epub 2014 Mar 24.
CRKL is recently defined as a new oncogene, which plays a role in the lung cancer progression. However, the potential mechanism of CRKL in human non-small cell lung cancer cell invasion is obscure. We investigated the potential mechanism of CRKL in lung cancer cell invasion using immunohistochemistry, plasmid transfection, Western blotting, real-time PCR, matrigel invasion assay, chromatin immunoprecipitation assay, and luciferase reporter assay. CRKL expression is higher in lymph node metastatic tumor compared with primary tumor. CRKL overexpression enhanced cell invasion and MMP9 expression in both HBE and H1299 cell lines. There was a significant correlation between CRKL overexpression and high MMP9 expression in primary tumors. MMP-9 antibody treatment significantly blocked cell invasion. CRKL overexpression also activated AP-1 luciferase reporter activity, ERK phosphorylation and association of c-fos to MMP9 promoter. Treatment with ERK inhibitor PD98059 in cells with CRKL transfection inhibited ERK activity, cell invasion, and MMP9 expression. These results suggested that overexpression of CRKL promoted cell invasion through upregulation of MMP9 expression and activation of ERK pathway.
CRKL最近被定义为一种新的癌基因，它在肺癌进展中发挥作用。然而，CRKL在人类非小细胞肺癌细胞侵袭中的潜在机制尚不清楚。我们使用免疫组织化学、质粒转染、蛋白质免疫印迹、实时聚合酶链反应、基质胶侵袭试验、染色质免疫沉淀试验和荧光素酶报告基因试验，研究了CRKL在肺癌细胞侵袭中的潜在机制。与原发性肿瘤相比，CRKL在淋巴结转移瘤中的表达更高。CRKL过表达增强了HBE和H1299细胞系中的细胞侵袭和MMP9表达。原发性肿瘤中CRKL过表达与高MMP9表达之间存在显著相关性。MMP-9抗体处理显著阻断细胞侵袭。CRKL过表达还激活了AP-1荧光素酶报告基因活性、ERK磷酸化以及c-fos与MMP9启动子的结合。在转染CRKL的细胞中用ERK抑制剂PD98059处理可抑制ERK活性、细胞侵袭和MMP9表达。这些结果表明，CRKL过表达通过上调MMP9表达和激活ERK途径促进细胞侵袭。
