The scaffold protein Tks5α is required for invadopodia-mediated cancer invasion both in vitro and in vivo. We have previously also revealed a role for Tks5 in tumor cell growth using three-dimensional (3D) culture model systems and mouse transplantation experiments. Here we use both 3D and high-density fibrillar collagen (HDFC) culture to demonstrate that native collagen-I, but not a form lacking the telopeptides, stimulated Tks5-dependent growth, which was dependent on the DDR collagen receptors. We used microenvironmental microarray (MEMA) technology to determine that laminin, fibronectin and tropoelastin also stimulated invadopodia formation. A Tks5α-specific monoclonal antibody revealed its expression both on microtubules and at invadopodia. High- and super-resolution microscopy of cells in and on collagen was then used to place Tks5α at the base of invadopodia, separated from much of the actin and cortactin, but coincident with both matrix metalloprotease and cathepsin proteolytic activity. Inhibition of the Src family kinases, cathepsins or metalloproteases all reduced invadopodia length but each had distinct effects on Tks5α localization. These studies highlight the crosstalk between invadopodia and extracellular matrix components, and reveal the invadopodium to be a spatially complex structure.

支架蛋白 Tks5α 是体外和体内侵袭伪足介导的癌症侵​​袭所必需的. 我们之前还使用三维 (3D) 培养模型系统和小鼠移植实验揭示了 Tks5 在肿瘤细胞生长中的作用。在这里，我们使用 3D 和高密度纤维状胶原 (HDFC) 培养来证明天然胶原-I，但不是缺乏端肽的形式，刺激依赖于 DDR 胶原受体的 Tks5 依赖性生长。我们使用微环境微阵列 (MEMA) 技术确定层粘连蛋白、纤连蛋白和原弹性蛋白也刺激了侵袭伪足的形成。Tks5α 特异性单克隆抗体显示其在微管和侵袭伪足上均有表达。然后使用胶原蛋白中和上的细胞的高分辨率和超分辨率显微镜将 Tks5α 放置在侵袭伪足的底部，与大部分肌动蛋白和皮质蛋白分开，但与基质金属蛋白酶和组织蛋白酶蛋白水解活性一致。Src 家族激酶、组织蛋白酶或金属蛋白酶的抑制都减少了侵袭伪足长度，但每一种对 Tks5α 定位都有不同的影响。这些研究突出了侵袭伪足和细胞外基质成分之间的串扰，并揭示了侵袭伪足是一种空间复杂的结构。