TDAG51 Deficiency Promotes Migration and Proliferation of Mouse Embryonic Fibroblasts.

Abstract T-cell death associated gene 51 (TDAG51) is a member of the pleckstrin homology-related domain family having pro-apoptotic characteristics. Indirect immunofluorescence studies demonstrate that endogenous TDAG51 co-localizes with focal adhesion kinase (FAK), a molecule found within focal adhesion complexes (FACs). FACs are believed to be the structural connection between the extracellular matrix (ECM) and actin cytoskeleton. Given that overexpression of TDAG51 promotes detachment-induced programmed cell death (PCD) and that TDAG51 co-localizes with FAK, it was proposed that deficiency of TDAG51 may stabilize FAC assembly, thereby affecting actin cytoskeletal organization and/or cell migration. To better understand the role of TDAG51 in cell migration, mouse embryonic fibroblasts (MEFs) deficient in TDAG51 were derived from TDAG51-deficient mice and compared to wild type MEFs. TDAG51-deficient MEFs showed increased migration following monolayer disruption or in response to chemotaxis on fibronectin-coated Boyden Chambers. In addition, loss of TDAG51 promotes the proliferation of MEFs. Collagen gel contraction experiments were performed to confirm the aggregate effect of the differences in migration and proliferation observed in TDAG51-deficient MEFs. Interestingly, migration and proliferation can be blocked by active b-integrin inhibitory antibody. In terms of a cellular phenotype, migratory TDAG51-deficient MEFs have distinct filopodial and lamellipodial extentions, compared to migratory wild type MEFs. Laser scanning confocal microscope demonstrated a significant decrease in F-actin stress fibers as well as increased distribution of vinculin within the lamellipodial protrusions of TDAG51-deficient MEFs. Importantly, reintroduction of TDAG51 into TDAG51-deficient MEFs reversed these phenotypic changes. Taken together, our findings suggest that loss of TDAG51 affects cell migration and proliferation through modifying focal adhesion complex assembly and cytoskeletal rearrangements.