Profilin1 regulates triple negative breast cancer cells migration through stabilization of Angiomotin and thereby YAP nuclear translocation

Abstract

The Hippo pathway effector YAP1 is a potent oncogenic driver in triple-negative breast cancer (TNBC) and its activity is restrained by the scaffold protein Angiomotin-p130 (AMOT). AMOT is itself short-lived, being targeted for proteasomal degradation by NEDD4-family E3 ubiquitin ligases that dock at its L/P-PxY motifs. Here we identify Profilin1 (PFN1), an actin-binding protein with established actin-independent tumour-suppressive signalling functions in TNBC as a direct binding partner and stabilizer of AMOT. PFN1 and AMOT are co-immunoprecipitated, they share 70 common interactors and NEDD4 is one of them. Protein-protein docking shows the interaction of PFN1 on the first PPxY motif of AMOT, through its actin-binding domain. We further show that PFN1s binding leaves the AMOT LPTY motif and both coiled-coil domains entirely unoccupied. Site-directed mutagenesis of AMOT PPxY motifs shows that PFN1 binding is unaffected by substitution of the PPxY tyrosines Y242 and Y287, either alone or in combination, indicating that PFN1 engages through its actin-binding domain. Functionally, PFN1 stabilizes AMOT as shown by cycloheximide-chase assay in TNBC. PFN1 induction increases cytoplasmic retention of YAP1, reduces TEAD occupancy at the CTGF promoter and thereby suppresses TNBC cell migration. Thus, this study suggests that PFN1 deregulates tumour cells migration by interacting with AMOT through its actin-binding domain, stabilizing AMOT and thereby arresting YAP in the cytoplasm, which might be an important therapeutic target to regulate TNBC.