| dc.description.abstract | The high incidence of invasive breast carcinomas, which accounts for about 90% of all breast cancer-related deaths, underscores the need for new and more effective therapeutic approaches to block the development of the malignant phenotype. Changes in the cytoskeletal organization of tumor cells and the physiochemical properties of the extracellular matrix (ECM) are key drivers of invasion and metastasis. Aberrant activation of phosphotyrosine signaling results in the formation of actin-based structures, called invadopodia, known to play an important role in ECM remodeling. We determined the role of Nck, adaptor proteins that link tyrosine phosphorylation with actin dynamics, in mammary carcinoma tumorigenesis and metastatic spread. We hypothesized that Nck-dependent actin remodeling promotes breast cancer invasion and metastasis by promoting the membrane accumulation/activity of MMP14, a key matrix-remodeling protease. To test this hypothesis, we used the highly metastatic MDA-MB-231 breast cancer cell line and a combination of three-dimensional (3D) tissue culture, optical imaging and a xenograft nude mouse model. To determine how Nck promotes invasion and metastasis by altering actin dynamics and membrane localization of MMP14 through regulation of Cdc42 activity we used gelatin degradation in conjunction with immunofluorescence microscopy, fluorescence recovery after photobleaching, total internal fluorescence microscopy and Förster resonance energy transfer.
The results show, for the first time, a requirement for Nck in breast carcinoma growth and invasion in 3D cultures and metastasis in vivo. Nck facilitates invasiveness by promoting invadopodia biogenesis and focalized matrix degradation. Loss of Nck results in decreased MMP14 accumulation at the ventral surface of invasive cells and altered actin dynamics at invadopodia. Loss of Nck also disrupted the localization of active Cdc42, a key regulator of trafficking and polarity. Our results support a model where Nck specifies patterns of Cdc42 activation and the targeted accumulation of MMP14 to facilitate the formation and maturation of invadopodia by integrating extracellular signals that modulate cytoskeletal dynamics. | en |
