Modeling M-phase control in Xenopus oocyte extracts: the surveillance mechanism for unreplicated DNA

Abstract

Alternating phases of DNA synthesis and mitosis, during the first 12 cell divisions of frog embryos, are driven by autonomous cytoplasmic oscillations of M-phase promoting factor (MPF). Cell-free extracts of frog eggs provide a convenient preparation for studying the molecular machinery that generates MPF oscillations and the surveillance mechanism that normally prevents entry into mitosis until chromosomal DNA is fully replicated. Early experiments suggested that unreplicated DNA blocks MPF activity by inducing phosphorylation of a crucial tyrosine residue, but recent evidence implicates a stoichiometric inhibitor (an MPF binding protein) as the 'braking' agent. Using a realistic mathematical model of the mitotic control system in frog egg extracts, we suggest that both tyrosine phosphorylation and a stoichiometric inhibitors are involved in the block of MPF by unreplicated DNA. Both pathways operate by raising the cyclin threshold for MPF activation. As a bonus, in the process of analyzing these experiments, we obtain more direct and reliable estimates of the rate constants in the model.

The dynamics of cell cycle regulation

“…In 1994-95, two experimental papers appeared, (58,59) reporting careful measurements of the rates of phosphorylation and dephosphorylation of Cdc2, Cdc25 and Wee1 in frog egg extracts in interphase and in M phase. From these data one can estimate directly the rate constants for these crucial steps in the model, (60) and the values come out very close to the original estimates of Novak and Tyson.…”

Regulation of the Eukaryotic Cell Cycle: Molecular Antagonism, Hysteresis, and Irreversible Transitions

“…By understanding the dynamical properties of restricted parts of the network "rst, we are able to put the pieces together into ever more comprehensive, computational models of intact control systems in speci"c organisms (Marlovits et al, 1998;Novak et al, 1998a, b;Novak & Tyson, 1997). In this paper we summarize what we have learned over the years about bistability and oscillations in the control system, and their relations to checkpoints and surveillance mechanisms in living cells.…”

“…At the core of the cell cycle is a hysteresis loop deriving from the fundamental antagonism between Cdk and APC (Novak et al, 1998) (0.90, 0.0045). Suppose a newborn cell resides at G1 (Cdh1 active and CycB missing).…”

Mathematical modeling as a tool for investigating cell cycle control networks

“…On the other hand, in an M-phase arrested extract (lanes a-c), tyrosine phosphorylation of Cdk1 by less active Wee1 is much slower, say V wee ′ ≈ 0.01 min −1 . Kumagai and Dunphy (1995) also show that the rate of tyrosine phosphorylation is 2.5 times slower when cycloheximide-treated extracts were supplemented with cyclin B monomers rather than with preformed cyclin B-Cdk1 dimers (their Figure 3, A and C), allowing Marlovits et al (1998) (43) to estimate that k 3 ≈ 0.005 nM −1 min −1 . Although only rough estimates, these values for V wee ′, V wee ″, and k 3 provide good starting points for more sophisticated parameter optimization procedures.…”