Cell cycle-regulated degradation of Xenopus cyclin B2 requires binding to p34cdc2.
The protein kinase activity of the cell cycle regulator p34cdc2 is inactivated when the mitotic cyclin to which it is bound is degraded. The amino (N)-terminus of mitotic cyclins includes a conserved "destruction box" sequence that is essential for degradation. Although the N-terminus of sea urchin cyclin B confer cell cycle-regulated degradation to a fusion protein, a truncated protein containing only the N-terminus of Xenopus cyclin B2, including the destruction box, is stable under conditions where full length molecules are degraded. In an attempt to identify regions of cyclin B2, other than the destruction box, involved in degradation, the stability of proteins encoded by C-terminal deletion mutants of cyclin B2 was examined in Xenopus egg extracts. Truncated cyclin with only the first 90 amino acids was stable, but other C-terminal deletions lacking between 14 and 187 amino acids were unstable and were degraded by a mechanism that was neither cell cycle regulated nor dependent upon the destruction box. None of the C-terminal deletion mutants bound p34cdc2. To investigate whether the binding of p34cdc2 is required for cell cycle-regulated degradation, the behavior of proteins encoded by a series of full length Xenopus cyclin B2 cDNA with point mutations in conserved amino acids in the p34cdc2-binding domain was examined. All of the point mutants failed to form stable complexes with p34cdc, and their degradation was markedly reduced compared to wild-type cyclin. Similar results were obtained when the mutant cyclins were synthesized in reticulocyte lysates and when cyclin mRNA was translated directly in a Xenopus egg extract. These results indicate that mutations that interfere with p34cdc2 binding also interfere with cyclin destruction, suggesting that p34cdc2 binding is required for the cell cycle-regulated destruction of Xenopus cyclin B2.
van der Velden
H M
HM
Department of Biological Sciences, University of Calgary, Canada.
Lohka
M J
MJ
D000818:Animals; D001483:Base Sequence; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D002453:Cell Cycle; D016213:Cyclins / Q000378:metabolism*; D008938:Mitosis; D008969:Molecular Sequence Data; D009865:Oocytes; D011485:Protein Binding; D017384:Sequence Deletion; D014982:Xenopus laevis / Q000378:metabolism
D016213:Cyclins; D016203:CDC2 Protein Kinase