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: In order to measure the activity of p53 in cells devoid of mDia2, we took advantage of shRNA and lentiviral infection because all tested siRNAs resulted in partial (less than 50%) and very transient down-regulation of mDia2 (supplemental Fig. S3C and unpublished observations). We obtained mDia2 knockdown cells having mDia2 protein levels reduced by about 75% compared with the control knockdown population (Fig. 5A). Silencing of mDia2 did not alter either the expression of FBXO3, HIPK2, p300, and p53 (Fig. 5A). The activity of p53 in 293T cells did not change upon mDia2 knockdown (not shown and Fig. 5D), thus suggesting that other Formins might cooperate with mDia2 in regulating p53.
Affiliation: From the ‡Division of Molecular Genetics, The Netherlands Cancer Institute, 1066 CX Amsterdam, The Netherlands;