Proteomic analyses uncover a new function and mode of action for mouse homolog of Diaphanous 2 (mDia2).
Bottom Line: Taking FBXO3 as a test case, we show that mDia2 binds FBXO3 and p53, and regulates p53 transcriptional activity in an actin-nucleation-independent and conformation-insensitive manner.Increased mDia2 and FBXO3 levels elevate p53 activity and expression thereby sensitizing cells to p53-dependent apoptosis, whereas their decrease produces opposite effects.Thus, we discover a new role of mDia2 in p53 regulation suggesting that the closed conformation is biologically active and an FBXO3-based mechanism to functionally specify mDia2's activity.
Affiliation: From the ‡Division of Molecular Genetics, The Netherlands Cancer Institute, 1066 CX Amsterdam, The Netherlands;Show MeSH
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Mentions: p53 is a tumor suppressor with a crucial function in the cellular apoptotic programs (41) and the interactome of wild-type mDia2 associates with the cell death functional group (Fig. 1B). Therefore, we monitored apoptosis in U2OS cells transfected with mDia2 and FBXO3, either alone or in combination, and compared it to the control ones. These experiments revealed that the activity of Caspase-3/7 was significantly increased in the cells expressing either mDia2 or FBXO3 compared with the control ones (Fig. 6A). Most importantly, the highest apoptotic index was measured in cells co-expressing mDia2 and FBXO3 (Fig. 6A). As elevated p53 levels were insufficient to trigger apoptosis in U2OS cells (Fig. 6A), there appears that mDia2 and FBXO3 jointly promote p53 activation. Consistent with this notion, (1) mDia2 and FBXO3 were present in the nuclear compartment both in control and etoposide-treated cells (supplemental Fig. S5), (2) knockdown of FBXO3, FMN1, and FMN1-mDia2 resulted in decreased p53 levels (Fig. 4D and Fig. 5A), whereas (3) co-expression of mDia2 and FBXO3 increased them (Fig. 4E).
Affiliation: From the ‡Division of Molecular Genetics, The Netherlands Cancer Institute, 1066 CX Amsterdam, The Netherlands;