Cell migration is essential for cellular morphogenesis and tissue repair, as well as a number of other important biological outcomes, and has a critical involvement in various developmental disorders and disease states. Of particular interest have been the regulatory events underlying cell migration as they relate to cell invasiveness, metastasis, and cancer progression. Tissue transglutaminase (tTG), a GTP- binding protein which also possesses an enzymatic transamidation activity that allows it to cross-link proteins, has been shown to play an important role in cell migration and invasion. As a signaling protein downstream of the epidermal growth factor (EGF) receptor, tTG becomes activated and is recruited to the leading edges of cells in response to EGF. In addition to its role in cell migratin and invasion, tTG has been shown to be important for Ras-driven transformation. The primary focus of this application is to learn more about how the activation and localization of tTG to the leading edges of migrating cells participates in the enhanced cell migration and invasive activity exhibited by human cancer cells and contributes to Ras- mediated cellular transformation. The following lines of investigation will be carried out to achieve this goal. 1) Determine the signalig mechanisms responsible for localizing tTG to the leading edges of actively migrating cells. EGF receptor signaling through Ras to JNK is important for tTG activation and localization.
This aim will involve identifying the substrates of JNK that mediate these effects. In particular, I will determine whether tTG or Hsp70 can serve as a substrate for JNK or whether other known substrates (i.e. paxillin) are important for regulating tTG activation and localization. 2) Determie how the chaperonin function of heat shock protein 70 (Hsp70) helps target tTG to the leading edges of cells. This line of study stems from our recent findings that Hsp70 and tTG interact and co-localize along the leading edges of cells and this localization is blocked in the presence of inhibitors of the ATP hydrolytic activity of Hsp70, myricetin, methylene blue, and VER 155008.
This aim will involve identifying the client proteins of Hsp70 which help target tTG to the leading edges of cells. 3) Determine how tTG works with Ras to promote cellular transformation. EGF signaling through Ras results in activation and localization of tTG to leading edges and Ras needs tTG for its transforming potential. In this aim, I will determine whether tTG- promoted Ras transformation occurs as a result of tTG's ability to enhance Ras activity or the activation of one of its downstream effectors, or whether tTG activates a Ras-independent signaling event that synergizes with Ras signaling to drive malignant transformation. These studies should provide insight into the significance of coupling tTG activation and localization to the leading edges of cells and how tTG is contributing to cell migration and invasion as well as cellular transformation.
Though many advances have been made in the development of cancer therapies throughout the years, one limitation is the fact that more advanced stage cancers often result in secondary metastases that are resistant to conventional treatment. For this reason, much effort is being devoted to understanding the molecular basis of enhanced cell migration and invasiveness as it pertains to cancer metastasis. In this study involving tissue transglutaminase, we hope to gain insight into the specific mechanisms used by cancer cells to enhance migration and invasive activity as this information is vital to the development of novel therapies for blocking cancer progression.