N-Terminal Basic Helix and S120 Phosphorylation Cooperatively Regulate Nuclear Localization of UBE2A/B in Mechanotransduction

Abstract

Simple Summary Cells experience physical cues from their environment, which influence gene expression through mechanotransduction. We previously found that Ubiquitin-conjugating enzyme E2 A/B (UBE2A/B) move into the nucleus under stiff or low-density conditions, promoting histone H2B (H2B) monoubiquitination and cell growth-related gene expression, but the underlying mechanism was unknown. Here, we identify key regulatory elements controlling UBE2A/B’s translocation. A phospho-mimetic mutation at S120 (S120D) enhances nuclear localization, while disruption of a basic N-terminal nuclear localization signal (3RA) reduces it. Cyclin-dependent kinase (CDK) inhibition blocks UBE2A/B nuclear entry and decreases H2B ubiquitination. We also show that the nuclear transport receptor Exportin-4 (XPO4) interacts specifically with phosphorylated UBE2A/B and that this interaction depends on both S120 phosphorylation and the nuclear localization signal (NLS) region. Structural modeling supports a cooperative binding mechanism. Together, our results reveal that UBE2A/B nuclear localization is controlled by phosphorylation-dependent signaling and NLS integrity, with XPO4 acting as a potential transport mediator.