The cell cycle is the carefully controlled sequence of events by which a cell duplicates its contents and divides into two daughter cells. Driving this progression is a family of enzymes known as Cyclin-Dependent Kinases (CDKs), with Cdc2 (also called CDK1) being the primary regulator for the transition into mitosis (M phase). Cdc2 activation requires the coordination of two distinct molecular events: physical partnership with a Cyclin protein and specific chemical modifications involving the addition and removal of phosphate groups. This regulatory system ensures the cell only enters division when all preparatory stages are complete.
Formation of the Cyclin-Cdc2 Complex
The first step in activating Cdc2 is forming a heterodimer complex with its regulatory partner, Cyclin B. Cdc2 is the catalytic subunit that transfers a phosphate group, while Cyclin B is the regulatory subunit that dictates when and where activity occurs. This association is necessary because the Cdc2 subunit alone is inactive, as a portion of its structure blocks the active site.
When Cyclin B binds, it causes a significant conformational change in the kinase molecule. This structural rearrangement partially exposes the active site by moving the flexible T-loop segment. Although this binding allows the enzyme to accept a substrate, the complex remains largely inactive until further modifications occur.
Essential Activating Phosphorylation at Threonine 161
The newly formed Cyclin B-Cdc2 complex requires an activating chemical modification to optimize its catalytic function. This modification is the phosphorylation of Threonine 161 (T161), located within the T-loop region of the Cdc2 molecule. This phosphorylation is carried out by the CDK-Activating Kinase (CAK), which is composed of CDK7 and Cyclin H.
The addition of the negatively charged phosphate group at T161 stabilizes the T-loop in an optimal conformation. This stabilization better aligns the active site’s catalytic residues, making the enzyme more efficient at binding ATP and target substrate proteins. While this T161 phosphorylation creates a potentially active enzyme, it is not yet fully functional because inhibitory signals are usually present.
Inhibitory Phosphorylation and the Primed State
Despite the activating signal from T161 phosphorylation, the Cyclin B-Cdc2 complex is held in an inactive or “primed” state during the G2 phase. This state is maintained by adding inhibitory phosphate groups to two sites near the ATP-binding pocket: Threonine 14 (T14) and Tyrosine 15 (Y15). These residues are positioned strategically to interfere with the kinase’s ability to bind ATP.
The addition of these inhibitory phosphates is the responsibility of specific kinases, primarily Wee1 and Myt1. Wee1 acts on Tyrosine 15, while Myt1 phosphorylates both T14 and Y15. The presence of these phosphate groups physically obstructs the ATP-binding site, overriding the activating effect of the T161 modification and pausing the cell cycle until preparation is complete.
Dephosphorylation Switch for Full Activation
The final transition into full Cdc2 kinase activity and the onset of mitosis is triggered by the rapid removal of the inhibitory phosphates. This process is governed by the dual-specificity phosphatase enzyme called Cdc25. Cdc25 specifically targets and removes the phosphate groups from both Threonine 14 and Tyrosine 15 on the Cdc2 subunit.
The removal of these two phosphates eliminates the physical obstruction in the ATP-binding pocket. This action causes a final conformational shift that results in the switch-like activation of the Cyclin B-Cdc2 complex. The newly active kinase then rapidly phosphorylates cellular targets, initiating structural changes associated with mitotic entry, such as nuclear envelope breakdown and chromosome condensation. Furthermore, the activated complex can itself phosphorylate and further activate Cdc25, establishing a positive feedback loop that ensures rapid commitment to mitosis.