Meyn Malcolm A, Melloy Patricia G, Li Jie, Holloway Sandra L
Howard Hughes Medical Institute and Department of Genetics, University of Pennsylvania Medical School, Philadelphia, PA 19104, USA.
Arch Biochem Biophys. 2002 Nov 15;407(2):189-95. doi: 10.1016/s0003-9861(02)00467-8.
Properly regulated cyclin proteolysis is critical for normal cell cycle progression. A nine-amino acid peptide motif called the destruction box (D box) is present at the N terminus of the yeast mitotic cyclins. This short sequence is required for cyclin ubiquitination and subsequent proteolysis. The anaphase-promoting complex/cyclosome (APC/C) is a multisubunit E3 required for cyclin ubiquitination. We have tested the D box of five mitotic cyclins for interaction with six APC/C subunits. The APC/C subunit Cdc23, but not five other subunits tested, interacted by two-hybrid analysis with the N terminus of wild-type Clb2. None of these subunits interacted with the N termini of the cyclins Clb1, Clb3, or Clb5. Mutations in the D box sequences of Clb2 inhibited interaction with Cdc23 both in vivo and in vitro. Our results provide the first evidence for a direct interaction between an APC/C substrate (Clb2) and an APC/C subunit (Cdc23).
细胞周期蛋白的适度调节性蛋白水解对于正常的细胞周期进程至关重要。酵母有丝分裂细胞周期蛋白的N端存在一个名为破坏框（D框）的九氨基酸肽基序。这个短序列是细胞周期蛋白泛素化及随后蛋白水解所必需的。后期促进复合物/细胞周期体（APC/C）是细胞周期蛋白泛素化所需的多亚基E3。我们测试了五种有丝分裂细胞周期蛋白的D框与六个APC/C亚基的相互作用。通过双杂交分析，APC/C亚基Cdc23与野生型Clb2的N端相互作用，而测试的其他五个亚基则没有。这些亚基均不与细胞周期蛋白Clb1、Clb3或Clb5的N端相互作用。Clb2的D框序列中的突变在体内和体外均抑制了与Cdc23的相互作用。我们的结果首次证明了APC/C底物（Clb2）与APC/C亚基（Cdc23）之间存在直接相互作用。
