A Role of TGFβ1 Dependent 14-3-3σ Phosphorylation at Ser69 and Ser74 in the Regulation of Gene Transcription, Stemness and Radioresistance

Transforming growth factor-beta (TGF beta) is a potent regulator of tumorigenesis, although mechanisms defining its tumor suppressing and tumor promoting activities are not understood. Here we describe phosphoproteome profiling of TGF beta signaling in mammary epithelial cells, and show that 60 identified TGF beta-regulated phosphoproteins form a network with scale-free characteristics. The network highlighted interactions, which may distribute signaling inputs to regulation of cell proliferation, metabolism, differentiation and cell organization. In this report, we identified two novel and TGF beta-dependent phosphorylation sites of 14-3-3 sigma, i.e. Ser69 and Ser74. We observed that 14-3-3 sigma phosphorylation is a feed-forward mechanism in TGF beta/Smad3-dependent transcription. TGF beta-dependent 14-3-3 sigma phosphorylation may provide a scaffold for the formation of the protein complexes which include Smad3 and p53 at the Smad3-specific CAGA element. Furthermore, breast tumor xenograft studies in mice and radiobiological assays showed that phosphorylation of 14-3-3 sigma at Ser69 and Ser74 is involved in regulation of cancer progenitor population and radioresistance in breast cancer MCF7 cells. Our data suggest that TGF beta-dependent phosphorylation of 14-3-3 sigma orchestrates a functional interaction of TGF beta/Smad3 with p53, plays a role in the maintenance of cancer stem cells and could provide a new potential target for intervention in breast cancer.