Apoptosis is critical for embryonic development, tissue homeostasis, and tumorigenesis and is determined largely by the Bcl-2 family of antiapoptotic and prosurvival regulators. Here, we report that glycogen synthase kinase 3 (GSK-3) was required for Mcl-1 degradation, and we identified a novel mechanism for proteasome-mediated Mcl-1 turnover in which GSK-3? associates with and phosphorylates Mcl-1 at one consensus motif (155STDG159SLPS163T; phosphorylation sites are in italics), which will lead to the association of Mcl-1 with the E3 ligase ?-TrCP, and ?-TrCP then facilitates the ubiquitination and degradation of phosphorylated Mcl-1. A variant of Mcl-1 (Mcl-1-3A), which abolishes the phosphorylations by GSK-3? and then cannot be ubiquitinated by ?-TrCP, is much more stable than wild-type Mcl-1 and able to block the proapoptotic function of GSK-3? and enhance chemoresistance. Our results indicate that the turnover of Mcl-1 by ?-TrCP is an essential mechanism for GSK-3?-induced apoptosis and contributes to GSK-3?-mediated tumor suppression and chemosensitization.