Phosphorylation-induced unfolding regulates p19INK4d during the human cell cycle.

Cell cycle progression is tightly regulated by cyclin-dependent kinases (CDKs). The ankyrin-repeat protein p19INK4d functions as a key regulator of G1/S transition; however, its molecular mode of action is unknown. Here, we combine cell and structural biology methods to unravel the mechanism by which p19INK4d controls cell cycle progression. We delineate how the stepwise phosphorylation of p19INK4d Ser66 and Ser76 by cell cycle-independent (p38) and -dependent protein kinases (CDK1), respectively, leads to local unfolding of the three N-terminal ankyrin repeats of p19INK4d This dissociates the CDK6-p19INK4d inhibitory complex and, thereby, activates CDK6. CDK6 triggers entry into S-phase, whereas p19INK4d is ubiquitinated and degraded. Our findings reveal how signaling-dependent p19INK4d unfolding contributes to the irreversibility of G1/S transition.