CCT Chaperonin Complex Is Required for the Biogenesis of Functional Plk1

Abstract

Experiments from several different organisms have demonstrated that polo-like kinases are involved in many aspects of mitosis and cytokinesis. Here, we provide evidence to show that Plk1 associates with chaperonin-containing TCP1 complex (CCT) both in vitro and in vivo. Silencing of CCT by use of RNA interference (RNAi) in mammalian cells inhibits cell proliferation, decreases cell viability, causes cell cycle arrest with 4N DNA content, and leads to apoptosis. Depletion of CCT in well-synchronized HeLa cells causes cell cycle arrest at G₂, as demonstrated by a low mitotic index and Cdc2 activity. Complete depletion of Plk1 in well-synchronized cells also leads to G₂ block, suggesting that misfolded Plk1 might be responsible for the failure of CCT-depleted cells to enter mitosis. Moreover, partial depletion of CCT or Plk1 leads to mitotic arrest. Finally, the CCT-depleted cells reenter the cell cycle upon reintroduction of the purified constitutively active form of Plk1, indicating that Plk1 might be a CCT substrate.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://mcb.asm.org/.

ACKNOWLEDGMENTS

We thank Eleanor Erikson and Xiaoyi Zhang for critical reading and comments on the manuscript and Ronald Melki for providing anti-CCTα antibody. We thank Kyung Lee and Toru Miki for providing HeLa cells stably expressing GFP-tubulin or H2B. We thank Hongtao Yu and Chou-Zen Giam for the Cdc20 construct and two anonymous reviewers for critical comments that improved the quality of the manuscript.

This work was supported by National Institutes of Health grant GM59172 to R.L.E. R.L.E. is the John F. Drum American Cancer Society Research Professor.