Maintaining the integrity of the ECM is critical in establishing a stringent balance between both pathological and physiological cellular events. During mammalian development, matrix turnover is vital to cell morphogenesis and tissue differentiation. Cells must be able to clear the barrier posed by the ECM in order to effectively respond to cues for tissue resorption, proliferation, and wound healing. Likewise, such degradation must be tightly regulated. Uncontrolled degradation of the ECM results in a wide array of degenerative diseases, while promoting tumor cell invasion and metastasis. Membrane Type-1 Matrix Metalloproteinase (MT1-MMP) has an essential role in matrix remodeling during physiological processes (Holmbeck et al. 1999; Zhou et al. 2000). Conversely, its enzymatic activity is key to acquiring a metastatic phenotype in a variety of tumor cells, including lung, colon, breast, and cervical carcinomas (Yana and Seiki 2002; Sabeh et al. 2004; Yana and Seiki 2003; Zhai et al. 2005; Itoh et al. 2006). The ability to alter the physical structure of the pericellular environment, while triggering the activation and modification of several cell surface proteins, identifies a central role for MT1-MMP in influencing cellular behavior. In return, the cell must be able to induce tight regulation of MT1-MMP enzymatic activity to prevent malignancy. The overall goal of this project was to investigate potential regulatory mechanisms that modulate MT1-MMP action at the cell surface.
Since its discovery, data examining the role of the cytoplasmic domain of MT1-MMP has been limited. Increasing evidence suggests that the cytoplasmic tail functions as a regulatory domain by its ability to mediate endocytosis, thereby limiting MT1-MMP surface expression (Uekita et al. 2001; Jiang et al. 2001). The mechanism underlying the ability of the tail to function in this capacity is unclear. In the current dissertation, data is presented that indicates MT1-MMP undergoes reversible phosphorylation at cytoplasmic residue threonine 567 (T567), and that this modification alters the functional activities of MT1-MMP when localized to the cell surface. The ability of the phospho-mimetic mutant (T567E) to enhance cell invasion, migration, and three-dimensional growth indicates that phosphorylation at this site may render the cell a heightened metastatic capacity.
An additional area of study has involved exploring cellular events that alter MT1-MMP trafficking. The Epidermal Growth Factor Receptor (EGFR) is overexpressed in 70% of ovarian tumors (Kohler et al. 1989; See et al. 2003), and as such, correlates with increased tumor progression and metastasis (Marmor et al. 2004; El-Rayes et al. 2004; Roskoski et al. 2004). Abrogation of EGFR signaling retards tumor progression (Alper et al. 2000; Brader et al. 1998), highlighting the importance of intracellular events mitigated by this receptor. An interesting link has been made between EGFR activation and the regulation of matrix degrading proteinases. In addition to elevated levels of EGFR, ovarian carcinomas display high levels of activated MT1-MMP, MMP-2, in addition to elevated levels of TIMP-2 (Davidson et al. 1999). Data presented here indicates that activation of the EGFR drives caveolae mediated endocytosis. This is of relevance because MT1-MMP preferentially localizes to caveolae (Annabi et al. 2001), placing EGFR and MT1-MMP in the same microdomain. Endocytosis is recognized as an efficient means of receptor attenuation in a variety of cell types. Removal of activated receptors from the cell surface abrogates the strength and duration of signal transduction, indicating a functional relationship between membrane trafficking and cell signaling. Internalization of MT1-MMP is recognized as a mechanism to regulate its surface expression. The ability of activated EGFR to facilitate such activity represents a novel mechanism by which EGFR regulates MT1-MMP trafficking from the cell membrane.
|Advisor:||Stack, Mary S.|
|Commitee:||Jones, Jonathan, Munshi, Hidyatullah, Rundell, Mary K., Vassar, Robert|
|Department:||Integrated Graduate Program in the Life Sciences|
|School Location:||United States -- Illinois|
|Source:||DAI-B 70/04, Dissertation Abstracts International|
|Keywords:||EGFR, Extracellular matrix, Matrix metalloproteinase|
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