SCF E3 ligase F-box protein complex SCFFBXL19 regulates cell migration by mediating Rac1 ubiquitination and degradation

摘要

Rac1, a member of the Rho family of GTPases, regulates diverse cellular functions, including cytoskeleton reorganization and cell migration. F-box proteins are major subunits within the Skp1-Cul1-F-box (SCF) E3 ubiquitin ligases that recognize specific substrates for ubiquitination. The role of F-box proteins in regulating Rac1 stability has not been studied. Mouse lung epithelial (MLE12) cells were used to investigate Rac1 stability and cell migration. Screening of an F-box protein library and in vitro ubiquitination assays identified FBXL19, a relatively new member of the F-box protein family that targets Rac1 for its polyubiquitination and proteasomal degradation. Overexpression of FBXL19 decreased both Rac1 active and inactive forms and significantly reduced cellular migration. Protein kinase AKT-mediated phosphorylation of Rac1 at serine⁷¹ was essential for FBXL19-mediated Rac1 ubiquitination and depletion. Lysine¹⁶⁶ within Rac1 was identified as a polyubiquitination acceptor site. Rac1^S71A and Rac1^K166R mutant proteins were resistant to FBXL19-mediated ubiquitination and degradation. Further, ectopically expressed FBXL19 reduced cell migration in Rac1-overexpressing cells (P<0.01, Rac1 cells vs. FBXL19+Rac1 cells), but not in Rac1 lysine¹⁶⁶ mutant-overexpressing cells. FBXL19 diminished formation of the migratory leading edge. Thus, SCF^FBXL19 targets Rac1 for its disposal, a process regulated by AKT. These findings provide the first evidence of an F-box protein targeting a small G protein for ubiquitination and degradation to modulate cell migration.—Zhao, J., Mialki, R. K., Wei, J., Coon, T. A., Zou, C., Chen, B. B., Mallampalli, R. K., Zhao, Y. SCF E3 ligase F-box protein complex SCF^FBXL19 regulates cell migration by mediating Rac1 ubiquitination and degradation.