👤 Patrick T Gunning

Focal adhesions (FAs) connect inner workings of cell to the extracellular matrix to control cell adhesion, migration and mechanosensing. Previous studies demonstrated that FAs contain three vertical layers, which connect extracellular matrix to the cytoskeleton. By using super-resolution iPALM microscopy, we identify two additional nanoscale layers within FAs, specified by actin filaments bound to tropomyosin isoforms Tpm1.6 and Tpm3.2. The Tpm1.6-actin filaments, beneath the previously identified α-actinin cross-linked actin filaments, appear critical for adhesion maturation and controlled cell motility, whereas the adjacent Tpm3.2-actin filament layer beneath seems to facilitate adhesion disassembly. Mechanistically, Tpm3.2 stabilizes ACF-7/MACF1 and KANK-family proteins at adhesions, and hence targets microtubule plus-ends to FAs to catalyse their disassembly. Tpm3.2 depletion leads to disorganized microtubule network, abnormally stable FAs, and defects in tail retraction during migration. Thus, FAs are composed of distinct actin filament layers, and each may have specific roles in coupling adhesions to the cytoskeleton, or in controlling adhesion dynamics. Show less

The Met receptor tyrosine kinase is activated or genetically amplified in some gastric cancers, but resistance to small-molecule inhibitors of Met often emerges in patients. We found that Met abundance correlated with a proliferation marker in patient gastric tumor sections, and gastric cancer cell lines that have MET amplifications depended on Met for proliferation and anchorage-independent growth in culture. Inhibition of Met induced temporal changes in gene expression in the cell lines, initiated by a rapid decrease in the expression of genes encoding transcription factors, followed by those encoding proteins involved in epithelial-mesenchymal transition, and finally those encoding cell cycle-related proteins. In the gastric cancer cell lines, microarray and chromatin immunoprecipitation analysis revealed considerable overlap between genes regulated in response to Met stimulation and those regulated by signal transducer and activator of transcription 3 (STAT3). The activity of STAT3, extracellular signal-regulated kinase (ERK), and the kinase Akt was decreased by Met inhibition, but only inhibitors of STAT3 were as effective as the Met inhibitor in decreasing tumor cell proliferation in culture and in xenografts, suggesting that STAT3 mediates the pro-proliferative program induced by Met. However, the phosphorylation of ERK increased after prolonged Met inhibition in culture, correlating with decreased abundance of the phosphatases DUSP4 and DUSP6, which inhibit ERK. Combined inhibition of Met and the mitogen-activated protein kinase kinase (MEK)-ERK pathway induced greater cell death in cultured gastric cancer cells than did either inhibitor alone. These findings indicate combination therapies that may counteract resistance to Met inhibitors. Show less