Tumor cell migration is a complex phenotype mediated by the genetic characteristics of the tumor cells in cooperation with the microenvironment. ADAM9, a member of a family of transmembrane matrix-metalloproteases, is implicated in cell-cell and cell-matrix interactions and the shedding of membrane receptors and ligands. ADAM9 is expressed as two alternatively spliced isoforms—membrane tethered ADAM9-Long (ADAM9-L) and secreted ADAM9-Short (ADAM9-S). ADAM9-L expression is observed in multiple epithelial cancers, while stromal cells in both the liver and bone secrete ADAM9-S. Expression of ADAM9-S increases breast cancer cell invasion, leading to the hypothesis that ADAM9 isoforms in breast tumors mediate tumor cell migration.
To evaluate the hypothesis that ADAM9-isoforms are functionally relevant for breast cancer progression, I first evaluated the expression of ADAM9 isoforms in breast cancer cell lines and tissues. Characterization of an ADAM9-S specific antibody allowed for the first confirmation of ADAM9-S expression in basal breast cancer cell lines and breast tumors. In contrast, ADAM9-L is expressed in all cell lines tested, regardless of subtype.
The remainder of my studies focused on the roles of ADAM9 isoforms in cell migration. Using Transwell assays to evaluate cell migration, I show that expression of ADAM9-S, but not ADAM9-L enhances breast cancer cell migration. A metalloprotease-deficient ADAM9-S is unable to increase migration, indicating that the function of ADAM9-S is metalloprotease-dependent. In contrast, shRNA studies reveal that ADAM9-L is a migration suppressor. Silencing both isoforms of ADAM9 using lentiviral-shRNA results in an increase in cell migration that is reversed by gene replacement of ADAM9-L, indicating that ADAM9-L suppresses cell migration. Subsequent studies of functional domain mutants reveal that ADAM9-L suppresses migration in a manner that is independent of metalloprotease activity, but requires the cytoplasmic domain of the protein. Integrin binding studies also reveal a putative role for the ADAM9-L integrin signaling, implicating ADAM9-L in a critical pathway mediating cell migration.
Together, these data show that ADAM9-L and ADAM9-S have opposing roles in cell migration. This finding has direct implications on future targeted therapy, and elucidates the need for isoform-specific study of other ADAMs.
|School Location:||United States -- Massachusetts|
|Source:||DAI-B 71/02, Dissertation Abstracts International|
|Subjects:||Molecular biology, Cellular biology, Pathology|
|Keywords:||Breast cancer, Cell migration, Metalloproteases|
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