The role of Tks adaptor proteins in invadopodia formation, growth and metastasis of melanoma

Abstract

Metastatic cancer cells are characterized by their ability to degrade and invade through extracellular matrix. We previously showed that the Tks adaptor proteins, Tks4 and Tks5, are required for invadopodia formation and/or function in Src-transformed fibroblasts and a number of human cancer cell types. In this study, we investigated the role of Tks adaptor proteins in melanoma cell invasion and metastasis. Knockdown of either Tks4 or Tks5 in both mouse and human melanoma cell lines resulted in a decreased ability to form invadopodia and degrade extracellular matrix. In addition, Tks-knockdown melanoma cells had decreased proliferation in a 3-dimensional type l collagen matrix, but not in 2-dimensional culture conditions. We also investigated the role of Tks proteins in melanoma progression in vivo using xenografts and experimental metastasis assays. Consistent with our in vitro results, reduction of Tks proteins markedly reduced subcutaneous melanoma growth as well as metastatic growth in the lung. We explored the clinical relevance of Tks protein expression in human melanoma specimens using a tissue microarray. Compared to non-malignant nevi, both Tks proteins were highly expressed in melanoma tissues. Moreover, metastatic melanoma cases showed higher expression of Tks5 than primary melanoma cases. Taken together, these findings suggest the importance of Tks adaptor proteins in melanoma growth and metastasis in vivo, likely via functional invadopodia formation.

Invadosomes are coming: new insights into function and disease relevance

“…Intriguingly, the Tks5‐KD tumors formed by both subcutaneous and tail vein injection displayed lower blood vessel density and blood vessel diameter when compared to controls, raising the possibility that tumor cell invadopodia may also play roles in tumor angiogenesis . In our breast cancer and melanoma studies, we also observed that cells with reduced Tks5 expression grew less well in matrices of type I collagen in vitro . It is of course possible that the reduced tumor growth observed in vitro and in vivo could be due to loss of invadopodia‐independent functions of Tks5.…”

“…)? We have shown in three different model systems (mouse fibrosarcoma, human breast cancer, and mouse and human melanoma) that knockdown of Tks5 reduces the in vivo growth of tumors, in subcutaneous, orthotopic, and metastatic sites . Intriguingly, the Tks5‐KD tumors formed by both subcutaneous and tail vein injection displayed lower blood vessel density and blood vessel diameter when compared to controls, raising the possibility that tumor cell invadopodia may also play roles in tumor angiogenesis .…”

“…Despite the similarities between Tks4 and Tks5, and the fact that Tks4 is also an important component of invadosome formation and function , care must be taken in ascribing all these developmental phenotypes to loss of podosomes. For example, recent research using a Tks4 knockout mouse revealed a differentiation defect in mesenchymal stromal cells (MSCs), caused by the reduced expression of RunX2 and Osterix, two osteogenic transcription factors that are normally up‐regulated during differentiation .…”

“…Three classes of proteases have been reported at invadosomes; zinc‐regulated matrix metalloproteases (e.g., MMP2, MMP9, MT1‐MMP, and the ADAMs family of sheddases), cathepsin cysteine proteases (e.g., cathepsin B); and serine proteases (e.g., seprase and urokinase‐type plasminogen activator, or uPA) . Of these, MT1‐MMP, a transmembrane MMP , has often been described as a master regulator of invadosome function . As well as ECM degradation and remodeling, pericellular proteases can function in the control of cell growth, apoptosis, and in cell–cell communications , through the release of growth factors that have a high affinity for matrix proteins (e.g., fibroblast growth factor, or FGF and transforming growth factor‐ β, or TGF‐β) , direct cleavage and activation of growth factors (e.g., TGF‐β and interleukin‐1 β) , and cleavage of cell surface receptors (e.g., FGF receptor 1) .…”

“…However, podosomes have also been observed in a number of other normal cell types, including osteoclasts , aortic endothelial cells , myoblasts , bronchial epithelial cells , neural crest cells , and in neuronal growth cones . Similarly, many cancer cell types form invadopodia, including pancreatic , ovarian , prostate , brain , skin , bladder , head and neck , breast , and melanoma .…”