Nck adapter proteins promote podosome biogenesis facilitating extracellular matrix degradation and cancer invasion

作者：Sankar P. Chaki, Rola Barhoumi, Gonzalo M. Rivera · 发表于：Cancer Medicine · 年份：2019 · DOI：10.1002/cam4.2640 · 被引用次数：24 · 研究领域：Wnt/β-catenin signaling in development and cancer、Microtubule and mitosis dynamics、Lymphatic System and Diseases

BACKGROUND: Podosomes are membrane-bound adhesive structures formed by actin remodeling. They are capable of extracellular matrix (ECM) degradation, which is a prerequisite for cancer cell invasion and metastasis. The signaling mechanism of podosome formation is still unknown in cancer. We previously reported that Nck adaptors regulate directional cell migration and endothelial lumen formation by actin remodeling, while deficiency of Nck reduces cancer metastasis. This study evaluated the role of Nck adaptors in podosome biogenesis and cancer invasion. METHODS: This study was conducted in vitro using both healthy cells (Human Umbilical Vein Endothelial Cell, 3T3 fibroblasts) and cancer cells (prostate cancer cell line; PC3, breast cancer cell line; MDA-MB-231). Confocal and TIRF imaging of cells expressing Green Fluorescence Protein (GFP) mutant under altered levels of Nck or downstream of kinase 1 (Dok1) was used to evaluate the podosome formation and fluorescent gelatin matrix degradation. Levels of Nck in human breast carcinoma tissue sections were detected by immune histochemistry using Nck polyclonal antibody. Biochemical interaction of Nck/Dok1 was detected in podosome forming cells using immune precipitation and far-western blotting. RESULTS: This study demonstrates that ectopic expression of Nck1 and Nck2 can induce the endothelial podosome formation in vitro. Nck silencing by short-hairpin RNA blocked podosome biogenesis and ECM degradation in cSrc-Y530F transformed ...