News and Publications

Control of Cell Division

S. Reed, B. Bertolaet, D. Clarke, S. Haase, J. Hanley-Hyde, L. Hengst, M. Henze, T. Herzinger, P. Kaiser, G. Mondesert, B. Niculescu, K. Sato, M. Segal, M. Smeets, C. Spruck, C. Tang, M. Watson, M. Wolfe, K. Won

Biological processes of great complexity can be approached by beginning with a systematic genetic analysis in which the relevant components are first identified and the consequences of their selective elimination by mutation are investigated. We have used yeast, which is uniquely tractable to this type of analysis, to investigate control of cell division. In recent years, it has become apparent that the most central cellular processes throughout the eukaryotic phylogeny are highly conserved in terms of both the regulatory mechanisms used and the proteins involved. Thus, it has been possible in many instances to generalize from yeast cells to human cells.

CONTROL IN YEAST

Most of our work in recent years has focused on the role and regulation of the Cdc28 protein kinase. Initially identified by means of a mutational analysis of the yeast cell cycle, this protein kinase is ubiquitous in eukaryotic cells and central to a number of aspects of control of cell-cycle progression.

Considerable progress has been made in understanding the regulation of the kinase in terms of it function at 2 points in the cell cycle: the transition from G₁ to S and the transition from G₂ to M. It appears that the same protein kinase catalytic subunit may perform these different functions by associating with other specific regulatory proteins known as cyclins to form active protein kinase complexes. Much of our recent effort has been devoted to identifying critical targets of these Cdc28 kinase activities. We have shown that many of the cell-cycle effects of Cdc28 kinase activity are mediated by regulating proteolysis of specific proteins.

We also found that a small Cdc28-associated protein, known as Cks1, appears to regulate the proteasome. Proteasomes are complex proteases that target ubiquitinated proteins, including important cell-cycle regulatory proteins. Our genetic analysis of the proteasome and ubiquitination pathways has yielded a complex web of interactions.

CONTROL IN MAMMALIAN CELLS

We have shown that the human homologs of the Cdc28 protein kinase are so highly conserved, structurally and functionally, relative to the yeast protein kinase, that they can function and be regulated properly in a yeast cell. Analyzing control of the cell cycle in mammalian cells, we produced evidence for the existence of regulatory schemes, similar to those elucidated in yeast, that use networks of both positive and negative regulators. We found that in human cells, agents that have positive or negative effects on proliferation exert these effects at the level of regulating cyclins and cyclin-dependent kinases. Of particular interest is the induction of cyclin-dependent kinase inhibitory proteins by antiproliferative signals. These proteins include not only members of the recently discovered Cip/Kip family of inhibitors but also p130, a relative of the retinoblastoma protein, implicated in human cancer.

As is the case in yeast, we have shown that proteolysis plays a key regulatory role in control of the cell in mammalian cells. We are determining if defects in proteolysis are implicated in human cancer. Finally, we are producing mice with defects in cell cycle--regulated proteolysis to observe the effect of such defects on development.

PUBLICATIONS

Dulic, V., Stein, G., Far, D.F., Reed, S.I. Nuclear accumulation of p21^Cip1/Waf1 at the onset of mitosis: A role at the G2/M-transition. Mol. Cell. Biol. 18:546, 1998.

Hengst, L., Lashuel, H.A., Reed, S.I. Complete inhibition of Cdk/cyclin by one molecule p21 Cip1. Genes Dev., in press.

Hengst, L., Reed, S.I. Inhibitors of the Cip/Kip family. Curr. Top. Microbiol. Immunol. 227:25, 1998.

Herzinger, T., Reed, S.I. Cyclin D3 is rate-limiting for the G1/S phase transition in fibroblasts. J. Biol. Chem., in press.

Kaiser, P., Sia, R.A.L., Bardes, E.G.S., Lew, D.J., Reed, S.I. Cdc34 and the F-box protein Met30 are required for degradation of the Cdk-inhibitory kinase Swe1. Genes Dev., in press.

Niculescu, A.B., Chen, X., Smeets, M., Hengst, L., Prives, C., Reed, S.I., Effects of p21^Cip1/Waf1 at both the G1/S and the G2/M cell cycle transitions: pRb is a critical determinant in blocking DNA replication and in preventing endoreduplication. Mol. Cell. Biol. 18:629, 1998.