Question

1. We learned about each of the following experimental stories in class, which together led to the discovery of cell cycle regulation. For each one, list the model organism used, the major experimental steps that were taken, and the major result that was obtained (the significant piece of cell cycle regulation that was uncovered by the experiment)

a. Identification of cyclin

b. Identification of cell division cycle (CDC) mutants

c. Identification of maturation promoting factor (MPF)


2. How did the purification of MPF in the late 1980s unite all three of the experimental stories listed in the previous question?

Answer 1

1)

a] Identification of cyclin- Scientist, Tim Hunt discovered the protein which seemed to control cell division, in sea urchin eggs. He radiolabelled the spawned sea urchin eggs to monitor patterns of protein synthesis after fertilization. When he observed the autoradiogram of the protein gel, he observed the a specific protein would accumulate in the cells and then disappear at cell division. He named this protein cyclin, on its property to form and then disappear in a cyclic manner. This discovery led to discovery of the fundamental mechanism determining cell division.

b] Identification of cell division cycle [cdc] mutants -- It was first identified in yeast, Saccharomyces cerevisiae.

Time lapse photomicroscopy has been utilized to detect temperature sensitive yeast mutants that are defective in gene functions needed at specific stages of cell-division cycle. Three genes, cdc-1, cdc-2 and cdc-3 execute early in the cell cycle but differ in their termination points. Cells carrying the cdc-1 mutation terminate at the bud formation point. Cells carrying the cdc-2 mutation terminate at mitosis. Cells carrying the cdc-3 mutation are defective in cell separation.

c] Identification of maturation promoting factor [MPF]- Maturation-promoting factor is the cyclin-Cdk complex that was discovered first in frog's eggs. It stimulates the mitotic and meiotic phases of the cell cycle. Cyclins and Cdks work together to control the cell cycle transitions through MPF. As M-cyclin accumulates, it binds to Cdks already present in the cell, forming complexes that are poised to trigger M phase.

So, MPF is a cell cycle checkpoint that regulates the passage of a cell from G2 growth phase to the M phase.

2) Maturation or M phase-promoting factor (MPF) is the universal inducer of M phase common to eukaryotic cells. MPF was originally defined as a transferable activity that can induce the G2/M phase transition in recipient cells. Today, however, MPF is assumed to describe an activity that exhibits its effect in donor cells, and furthermore, MPF is consistently equated with the kinase cyclin B-Cdk1. In some conditions, however, MPF, as originally defined, is undetectable even though cyclin B-Cdk1 is fully active. For over three decades, this inconsistency has remained a long-standing puzzle. The enigma is now resolved through the elucidation that MPF, defined as an activity that exhibits its effect in recipient cells, consists of at least two separate kinases, cyclin B-Cdk1 and Greatwall (Gwl). Involvement of Gwl in MPF can be explained by its contribution to the autoregulatory activation of cyclin B-Cdk1 and by its stabilization of phosphorylations on cyclin B-Cdk1 substrates, both of which are essential when MPF induces the G2/M phase transition in recipient cells. To accomplish these tasks, Gwl helps cyclin B-Cdk1 by suppressing protein phosphatase 2A (PP2A)-B55 that counteracts cyclin B-Cdk1. MPF, as originally defined, is thus not synonymous with cyclin B-Cdk1, but is instead a system consisting of both cyclin B-Cdk1 that directs mitotic entry and Gwl that suppresses the anti-cyclin B-Cdk1 phosphatase. The current view that MPF is a synonym for cyclin B-Cdk1 in donor cells is thus imprecise; instead, MPF is best regarded as the entire pathway involved in the autoregulatory activation of cyclin B-Cdk1, with specifics depending on the experimental system.