Cell Cycle Checkpoints and Related Concepts

Course Overview and Reminders

This section includes comprehensive information about upcoming assignments and labs for the semester, detailing the structure of remaining classes and exams, as well as essential dates and deadlines to ensure all students are well-informed.

Upcoming Lab Assignments

  • In Search of the Cancer Gene Lab: This is the last required lab of the semester, providing hands-on experience in molecular biology techniques to identify and analyze genetic mutations implicated in various cancers. It has been passed out for the current week, and its completion is crucial for fulfilling course requirements.

Class Schedule and Important Dates

  • Thanksgiving Break: No classes will be held on Wednesday of that week, consequently leading to no scheduled lab sessions. Students should plan accordingly.

  • Makeup Lab: A dedicated makeup session is scheduled for the last week of the semester, specifically on December 3. This session is designed for students who may have missed a required lab for legitimate reasons and need to complete it. If a student has attended and completed all regular labs, they do not need to attend this makeup session, as its primary purpose is to ensure all students meet the lab completion requirement.

  • Perusal Assignments:

    • Another perusal assignment is due tomorrow night. These assignments are designed to encourage active reading and understanding of course material before lectures, often involving critical thinking questions on key concepts from readings.

    • Students should be aware of a perusal assignment due on Sunday night during Thanksgiving. This will be available on Canvas well in advance and can be completed prior to the break to avoid last-minute stress. The absolute completion deadline for this assignment is November 30.

  • Final Exam and OOPS Tokens:

    • The final exam date is December 10. During finals week, OOPS tokens, which provide students with the opportunity for extensions or make-up work without penalty, must be redeemed by December 9, the day before the final exam. This deadline is strict to ensure all grading and academic records can be finalized promptly.

Overview of Upcoming Topics and Content

  • Next class (Wednesday): Discussion on apoptosis and p53. Apoptosis is programmed cell death, a crucial process for development and eliminating damaged or unwanted cells. p53 is a tumor suppressor protein, often called 'the guardian of the genome', playing a central role in initiating apoptosis or cell cycle arrest in response to cellular stress, especially DNA damage.

  • Review of the sixth exam, a week from today.

  • The last week of the semester will focus extensively on the hallmarks of cancer, culminating in comprehensive discussions that integrate and synthesize knowledge accumulated throughout the semester regarding cancer biology.

Recap of Previous Lecture on the Cell Cycle Checkpoints
Key Topics Discussed

  • Cell Cycle Checkpoints: Their roles as critical regulatory junctures in the eukaryotic cell cycle, monitoring internal and external conditions to ensure proper cell division, preventing the propagation of damaged cells or incomplete replication, and thereby maintaining genomic integrity. The proteins regulating progression through these checkpoints were emphasized.

G2 to M Checkpoint

  • Purpose: At this checkpoint, the cell meticulously checks for adequate size, assesses the integrity of its genome for any DNA damage, and confirms that all DNA replication has been accurately completed before proceeding to mitosis. This prevents the division of cells with mutated or incompletely replicated DNA.

  • Checks:

    • Cell Size: The cell must meet specific growth requirements, ensuring it has sufficient biomass and cellular components to divide into two viable daughter cells.

    • DNA Damage: If DNA damage is detected, the checkpoint pauses the cell cycle, activating repair mechanisms to fix the damage. This allows time for repair before the DNA is divided among daughter cells, preventing the transmission of mutations.

    • DNA Replication: Ensures that all chromosomes have been completely and accurately replicated, avoiding the distribution of incomplete genetic material to daughter cells.

Regulatory Proteins Involved

  • Cyclin Dependent Kinase (CDK): These are serine/threonine protein kinases that are constitutively present throughout the cell cycle but are inactive on their own. Their activity is entirely dependent on binding to cyclins.

  • Cyclins: These proteins vary in concentration during the cell cycle, being synthesized and subsequently degraded at specific phases. Cyclins bind to and activate CDKs, inducing conformational changes that expose the CDK's active site. Their degradation and activity are crucial for allowing the cell to transition smoothly between different cell cycle phases. Different cyclins (e.g., G1 cyclins, S cyclins, M cyclins) lead to the phosphorylation of distinct target proteins, guiding the cell toward various stages (e.g., S phase and mitosis).

Checkpoints in Mitosis

  • Between metaphase and anaphase: This critical control point is known as the Spindle Assembly Checkpoint (SAC) or metaphase checkpoint. It rigorously ensures that all sister chromatids are correctly bioriented – meaning each chromatid is fully and stably attached to kinetochore microtubules originating from opposite spindle poles. This ensures accurate chromosome segregation and prevents aneuploidy.

    • APC (Anaphase Promoting Complex): The APC/C (Anaphase Promoting Complex/Cyclosome) is an E3 ubiquitin ligase that activates this checkpoint by adding ubiquitin chains to specific target proteins for degradation by the 26S proteasome, rather than phosphorylating them. This ubiquitin-mediated proteolytic event is a key method for advancement out of metaphase.

    • It is critical in degrading mitotic cyclins (e.g., Cyclin B) which allows cells to inactivate mitotic CDKs and enter telephase and cytokinesis.

    • Cohesins: These are protein complexes (composed of Smc1, Smc3, Scc1/Rad21, Scc3 subunits) that encircle sister chromatids, holding them together after DNA replication until anaphase. This cohesion is vital for proper segregation. Separase, an enzyme responsible for cleaving cohesins, is kept inactive by being bound to the inhibitory protein securin. Separase is activated by the APC complex via the degradation of securin.

Transition from Metaphase to Anaphase

  • The APC complex tags securin with ubiquitin chains, marking it for degradation by the proteasome. Degradation of securin releases and activates separase, which then cleaves the Scc1 subunit of cohesins.

  • Without cohesins holding them together, sister chromatids can be pulled apart by the retracting spindle fibers, moving towards opposite poles during anaphase, thereby ensuring equal distribution of genetic material.

The G1 to S Checkpoint

  • This is a critical decision point where the cell commits to DNA replication. It scrutinizes the external environment for the presence of sufficient growth factors to stimulate division, adequate nutrients for biosynthesis, and whether the cell has attained a sufficient size. Importantly, it also checks for any DNA damage before replication begins, pausing the cycle if factors are absent or damage is present.

  • Regulated by a G1-S cyclin CDK complex (e.g., Cyclin D-CDK4/6 and Cyclin E-CDK2):

    • E2F: This is a transcription factor responsible for initiating the gene expression program required for DNA replication. Its activation is normally inhibited by Rb (retinoblastoma) protein, a tumor suppressor. The G1-S cyclin CDK complex phosphorylates Rb, causing it to undergo a conformational change and release E2F. Free E2F then activates the transcription of S-phase genes (e.g., DNA polymerase, dihydrofolate reductase), committing the cell to DNA synthesis.

Role of Growth Factors in Progression

  • Growth factors trigger specific external cell signaling pathways that activate the cyclin-CDK complexes necessary for progression through the cell cycle. This often involves receptor-ligand interactions at the cell surface.

  • The discussion included the RAS signaling pathway, which is a key cell communication system that transmits signals from outside the cell to the nucleus, regulating cell growth, proliferation, differentiation, and survival. It typically starts with growth factor binding to a receptor tyrosine kinase (RTK), leading to the activation of the small G protein RAS. Activated RAS then triggers a multi-step phosphorylation cascade (the MAPK/ERK pathway, involving RAF-MEK-ERK), which ultimately leads to the activation of various transcription factors that promote the expression of genes necessary for cell cycle progression and, specifically, the synthesis of G1 cyclins.

Next Steps and Videos

  • A forthcoming video will demonstrate the intricate relationships between the discussed proteins, signaling pathways, and interaction mechanisms, clarifying the direct connections between growth factors and the regulation of cyclins.

  • A detailed discussion and elaboration of p53 functions within cell signaling control and checkpoints will occur in the next class, particularly focusing on its extensive connections to the initiation of apoptosis and its critical role as a tumor suppressor.