Maturation of Human Cyclin E Requires the Function of Eukaryotic Chaperonin CCT

ABSTRACT

Cyclin E, a partner of the cyclin-dependent kinase Cdk2, has been implicated in positive control of the G₁/S phase transition. Whereas degradation of cyclin E has been shown to be exquisitely regulated by ubiquitination and proteasomal action, little is known about posttranscriptional aspects of its biogenesis. In a yeast-based screen designed to identify human proteins that interact with human cyclin E, we identified components of the eukaryotic cytosolic chaperonin CCT. We found that the endogenous CCT complex in yeast was essential for the maturation of cyclin E in vivo. Under conditions of impaired CCT function, cyclin E failed to accumulate. Furthermore, newly translated cyclin E, both in vitro in reticulocyte lysate and in vivo in human cells in culture, is efficiently bound and processed by the CCT. In vitro, in the presence of ATP, the bound protein is folded and released in order to become associated with Cdk2. Thus, both the acquisition of the native state and turnover of cyclin E involve ATP-dependent processes mediated by large oligomeric assemblies.

ACKNOWLEDGMENTS

We thank John Cogswell and Sue Neill of Glaxo Wellcome for providing the recombinant adenoviruses and Suwon Kim for the CCT mutant yeast strains. We thank members of the Reed laboratory and the Horwich laboratory for valuable discussions.

This work was supported by U.S. Army grant DAMD 17-94-J4208 to S.I.R. and National Institutes of Health Postdoctoral Fellowship CA09292 to K.-A.W. A.L.H. is an investigator of the Howard Hughes Medical Institute.