The cell cycle is one of the most crucial and complex processes in living organisms, responsible for regulating cell growth, division, and replication, thereby maintaining normal development and tissue repair. In eukaryotes, the progression of the cell cycle is driven by a series of precisely regulated events, including protein phosphorylation, dephosphorylation, and the formation and dissociation of protein complexes. Among these, Cyclin-Dependent Kinases (CDKs) are the core elements of cell cycle regulation. By forming complexes with cyclins, CDKs drive the transition of cells from one phase of the cell cycle to the next. This article provides an in-depth exploration of the molecular structure, functional mechanisms, roles in the cell cycle, and disease associations of the CDK family.

Historical Background and Discovery of the CDK Family

The discovery of CDKs is closely linked to the development of cell cycle research. In the 1980s, researchers first identified a gene in yeast, CDC28, that was closely related to cell cycle progression. Subsequently, scientists discovered that the homolog of CDC28 in fission yeast is CDC2, revealing the crucial role these genes play in cell cycle regulation. Further studies demonstrated that CDC28 and CDC2 encode serine/threonine protein kinases, which regulate cell cycle progression by forming complexes with periodically expressed cyclins.

Building on yeast research, scientists further identified multiple CDK members in more complex eukaryotes and found that these CDKs play critical roles at various stages of the cell cycle. These discoveries laid the theoretical foundation for understanding cell cycle regulation and provided important targets for subsequent cancer research and drug development.

Classification and Functions of the CDK Family

Members of the CDK family are highly conserved in terms of function and structure, but their spatiotemporal specificity is critically important. Based on functional differences, the CDK family can be broadly divided into two categories: cell cycle CDKs and transcriptional regulatory CDKs.

1. Cell Cycle CDKs: These CDKs are directly involved in the regulation of the cell cycle, including CDK1, CDK2, CDK4, and CDK6.

CDK1: Also known as CDC2, CDK1 is a key kinase in cell cycle progression. It regulates the transition from the G2 phase to the M phase by forming complexes with cyclins A and B. The activity of CDK1 is influenced by various regulatory factors, such as precise control of activating and inactivating phosphorylation events.

CDK2: Primarily functions during the G1/S transition and S phase by forming complexes with cyclins E and A, initiating DNA replication, and ensuring smooth progression of the cell cycle. Aberrant CDK2 activity is often associated with tumorigenesis.

CDK4/6: These CDKs regulate cell cycle progression during the G1 phase by forming complexes with cyclin D, particularly controlling the passage through the G1/S checkpoint. Overactivation of CDK4/6 is commonly associated with various cancer types.

2. Transcriptional Regulatory CDKs: These CDKs mainly participate in the regulation of transcription, including CDK7, CDK8, CDK9, and others.

CDK7: Functions as a CDK-activating kinase (CAK), not only activating other cell cycle CDKs through its association with cyclin H but also phosphorylating RNA polymerase II, thereby regulating gene transcription.

CDK8/19: Associates with the Mediator complex, involved in the regulation of transcription. Dysregulation of CDK8 is linked to transcriptional disorders in various cancers.

CDK9: Regulates transcriptional elongation by associating with cyclin T, making it a critical node in gene transcription regulation.

Figure 1. Evolutionary relationships among mammalian CDK subfamilies. (Malumbres M. et al., 2014)

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Molecular Structure and Activation Mechanisms of CDKs

CDK family members exhibit highly conserved structural features, mainly comprising three functional domains: the N-terminal kinase domain rich in helices, a connecting region, and a C-terminal ATP-binding pocket. The activity of CDKs primarily depends on conformational changes in the ATP-binding pocket and the activation phosphorylation following the binding of CDKs to cyclins.

1. Cyclin Binding: The activity of CDKs depends on their binding to cyclins. The expression and degradation of cyclins are central to cell cycle regulation. At different stages of the cell cycle, specific cyclins accumulate in the cell and bind to the corresponding CDKs to form active complexes. For example, CDK1 forms the MPF complex with cyclin B during the M phase, initiating mitosis.

2. Activating Phosphorylation: Full activation of CDKs requires phosphorylation at specific amino acid residues. CDK7, functioning as CAK, activates CDKs by phosphorylating the Thr160 residue (or equivalent sites) on CDKs. Additionally, CDKs can be further regulated by dephosphorylation by phosphatases such as CDC25, which removes inhibitory phosphorylation, thereby fine-tuning CDK activity.

3. Regulation by CKIs: Cyclin-dependent kinase inhibitors (CKIs) are key negative regulators of CDK activity. CKIs inhibit CDK activity by directly binding to CDKs or CDK-cyclin complexes. For instance, p21 and p27 are classic CKIs that bind to the CDK2-cyclin E complex, preventing the cell from entering the S phase, and thereby acting as a "brake" in the cell cycle.

Spatiotemporal Regulation of the Cell Cycle by the CDK Family

The CDK family drives the smooth progression of the cell cycle from the G1 phase to the M phase by periodically binding to different cyclins. This regulation exhibits high spatiotemporal specificity:

1. Transition from G1 to S Phase: In the early G1 phase, CDK4 and CDK6 bind to cyclin D, initiating the phosphorylation of Rb protein and releasing the E2F transcription factor, thereby promoting the expression of G1/S phase genes. As the cell progresses into the late G1 phase, CDK2 binds to cyclin E, further pushing the cell past the G1/S checkpoint and initiating DNA replication.

2. Progression from S Phase to G2 Phase: During the S phase, CDK2 binds to cyclin A to regulate the initiation and progression of DNA replication, ensuring genomic integrity and stability. Subsequently, CDK2 activity gradually decreases, and the CDK1-cyclin A complex starts functioning in the G2 phase, preparing the cell for mitosis.

3. Entry from G2 to M Phase: Cells in the G2 phase need to pass a series of checkpoints to ensure that DNA replication is complete and undamaged. CDK1 binds to cyclin B to form the MPF complex, driving the cell into the M phase and initiating nuclear envelope breakdown and chromosome condensation. The activation of MPF is the key step of the M phase, and the balance between its activation and inhibition determines whether the cell can successfully enter mitosis.

4. Completion of M Phase and Cell Division: In the late M phase, the APC/C (Anaphase-Promoting Complex/Cyclosome) mediates the degradation of cyclin B, leading to MPF inactivation, thereby initiating the completion of mitosis. The inactivation of CDK1 and the degradation of cyclins ensure that the cell can complete division and return to the G1 phase.

CDKs and Cell Cycle Dysregulation in Cancer

Aberrant regulation of the cell cycle is closely associated with the development of various diseases, particularly cancer. In many types of cancer, abnormalities in CDKs and their regulatory mechanisms are the root cause of uncontrolled cell proliferation.

1. CDK4/6 and Breast Cancer: In breast cancer, aberrant CDK4/6 activity is often associated with dysregulated cell cycle control. CDK4/6, through binding with cyclin D, drives cells through the G1/S checkpoint, thereby promoting cancer cell proliferation. CDK4/6 inhibitors, such as Palbociclib, have been widely used in the treatment of breast cancer. These inhibitors prevent cell cycle progression by inhibiting CDK4/6 activity, thereby suppressing tumor growth.

2. CDK2 and Ovarian Cancer: Overexpression or increased activity of CDK2 is common in various tumors, including ovarian cancer. CDK2 plays a crucial role in the G1/S transition, and its aberrant activity leads to uncontrolled DNA replication and cell proliferation. Therefore, targeting CDK2 is increasingly being considered as a promising approach in anticancer therapy.

3. CDK9 and Leukemia: CDK9 regulates transcriptional elongation and plays a critical role in leukemia. Abnormal activity of CDK9 in leukemia cells often leads to the overexpression of anti-apoptotic genes, thereby promoting cancer cell survival. Inhibiting CDK9 is considered a potential anti-leukemia strategy, effectively suppressing cancer cell growth by targeting CDK9 activity.

Clinical Applications and Prospects of CDK Inhibitors

As understanding of the critical role of CDKs in cancer and other diseases deepens, CDK inhibitors have emerged as an important part of targeted therapies. Several CDK inhibitors have been applied clinically or are under development.

1. CDK4/6 Inhibitors: Drugs such as Palbociclib, Ribociclib, and Abemaciclib selectively inhibit the activity of CDK4/6, blocking cancer cells from progressing through the G1/S phase and exerting anticancer effects. These drugs have shown significant efficacy in breast cancer treatment and have become a first-line therapy for hormone receptor-positive breast cancer.

2. Pan-CDK Inhibitors: Drugs such as Flavopiridol inhibit multiple CDK family members, including CDK1, CDK2, and CDK9, exhibiting broad-spectrum antitumor activity. However, their clinical application is limited due to side effects resulting from non-selective inhibition.

3. CDK9 Inhibitors: Drugs such as BAY 1143572 target CDK9 and inhibit transcriptional elongation, thereby suppressing cancer cell growth. Research into CDK9 inhibitors in hematologic malignancies is gaining attention, showing promising clinical prospects.

The CDK family plays a central role in cell cycle regulation, and studies on its molecular mechanisms have not only elucidated fundamental aspects of cell proliferation but also provided critical targets for the treatment of cancer and other diseases. As research progresses, more CDK family members and their regulatory mechanisms will be discovered, opening new avenues for disease diagnosis and treatment. The development and application of CDK inhibitors will continue to advance, laying the foundation for the era of precision medicine. In the future, exploring the molecular mechanisms linking the CDK family to diseases will help develop more effective therapeutic strategies, ultimately benefiting more patients.