The KEN box regulates Clb2 proteolysis in G1 and at the metaphase-to-anaphase transition

Clb2 mitotic cyclin inhibits cell cycle progression by preventing mitotic e xit and DNA synthesis. To allow cell cycle progression, Clb2 proteolysis is triggered by Cdc20 during the meta phase-to-ana phase (M-A) transition and by Hct1 during mitotic exit and G1 [1-6]. A cis element called the destruc tion box is required for this proteolysis [7-11]. Recently, an additional c is element called the "KEN box" was also shown to be required for proteolys is of human CDC20 and Securin [3, 12]. Using a novel color assay, we show t hat a Clb2 KEN box is required to target a fusion protein containing the fi rst 124 amino acids of Clb2 for proteolysis. We further show that full-leng th Clb2 bearing mutations in the KEN box is degraded efficiently during the M-A transition, but poorly during G1. If the destruction box of Clb2 is mu tated in combination with mutation of the KEN box, then this form of Clb2 i s more stable than Clb2 bearing either mutation by itself during both M-A a nd G1. Our results show that the KEN box and the destruction box act togeth er during both M-A and G1 to regulate Clb2 proteolysis.