Tissue Transglutaminase And Its Effects On Cell Migration And Survival

Abstract

Tissue transglutaminase (tTG) is a GTPase and acyl transferase which catalyzes the formation of covalent crosslinks between two protein substrates. tTG expression and activation are frequently up-regulated in different types of human cancer, where it has been shown to be important for enhancing cell motility. Using HeLa cervical carcinoma cells as a model system, I show that a membrane-associated pool of tTG becomes activated and re-distributes to the leading edges of migrating cells upon EGF stimulation. Immunoprecipitations of tTG from the membrane fractions of EGF-treated HeLa cells led to the discovery that tTG binds the heat shock protein (Hsp)70 family of molecular chaperones. tTG and Hsp70 co-localize at leading edges, and this localization is dependent on the ATP hydrolytic activity of Hsp70, as inhibitors against this function prevent both tTG and Hsp70 from localizing to leading edges. More importantly, these inhibitors also block the EGF-dependent migration of HeLa cells and the constitutive migration of MDA-MB231 cells, suggesting that Hsp70 helps localize tTG to leading edges to facilitate its role in promoting cell migration. While tTG has been shown to influence a number of aspects of cancer progression, to what degree tTG works with oncogenic proteins to elicit these outcomes versus its intrinsic ability to impact malignant transformation is unknown. Thus, I have examined how ectopic expression of tTG in a normal (non-transformed) cellular context influences the behavior of these cells. Using NIH3T3 fibroblasts stably expressing the vector alone or a Myc-tagged form of wild-type tTG, I found that tTG strongly protected these cells from serum-starvation-induced apoptosis by activating the PI3-kinase/mTOR/p70 S6-kinase pathway. tTG binds c-Src and PI3kinase, and the formation of this complex is critical for the activation of the PI3-kinase signaling pathway. Activation of PI3-kinase signaling is essential for tTG's ability to promote cell survival, as inhibition of any component in this pathway, including Src, PI3-kinase, or mTOR, eliminates the protective effect afforded to the cells by tTG expression.