Cell Cycle Notes
The Cell Cycle
Cell division is essential for reproduction and survival of all living organisms.
The cell cycle involves duplication of cell contents and division in two.
In unicellular organisms, cell division creates a new organism.
In multicellular organisms, it's necessary for development and replacing dead cells.
Stopping cell division entirely would lead to death within days.
The cell cycle varies among organisms, but the core goal is to pass on genetic information.
DNA in chromosomes is replicated to produce identical copies.
Replicated chromosomes are segregated into daughter cells, each receiving a copy of the genome.
Cells also duplicate organelles and macromolecules to maintain size.
Cell growth (increase in mass) is coordinated with cell division.
Overview of the Cell Cycle
The cell cycle's primary function is DNA duplication and segregation into two identical daughter cells.
This occurs in two major phases:
S phase (DNA synthesis):
Requires 10-12 hours in mammalian cells, about half the cycle time.
M phase (mitosis):
Requires less time (less than an hour in mammalian cells).
Involves nuclear division (mitosis) and cytoplasmic division (cytokinesis).
Sister chromatids:
DNA molecules in duplicated chromosomes remain intertwined after S phase.
In prophase (early mitosis), they condense into compact rods.
Sister-chromatid cohesion keeps them linked.
Attached to mitotic spindle after the nuclear envelope disassembles.
Metaphase:
Sister chromatids align at the spindle equator.
Anaphase:
Sister-chromatid cohesion is destroyed.
Sister chromatids are pulled to opposite spindle poles.
Telophase:
Spindle disassembles, chromosomes packaged into separate nuclei.
Cytokinesis:
Cleaves the cell in two, each daughter cell inheriting one nucleus.
Eukaryotic Cell Cycle Phases
Eukaryotic cell cycle has four phases: G1, S, G2, and M.
Gap phases (G1 and G2) provide time for cell growth and monitoring conditions.
G1 phase is critical for monitoring external environment.
Unfavorable conditions lead to cell cycle delay or Go resting state.
Cells in Go can remain for extended periods (days, weeks, years).
Favorable conditions or signals trigger progression through Start (yeasts) or restriction point (mammalian cells).
After Start, cells are committed to DNA replication.
Variations in cell cycle:
Early vertebrate embryo divisions lack growth, alternating S and M phases.
Endocycle (endoreduplication) involves multiple S phases without M phases, leading to increased gene product production.
Mitosis without cytokinesis results in multinucleated cells.
Cell-Cycle Control in Eukaryotes
The basic organization of the cell cycle is conserved in all eukaryotic cells.
Eukaryotes use similar machinery and control mechanisms.
Proteins of the cell-cycle control system appeared over a billion years ago and are highly conserved.
Model organisms used in cell-cycle analysis:
Budding yeast (Saccharomyces cerevisiae) and fission yeast (Schizosaccharomyces pombe): Genetic and molecular approaches.
Frog (Xenopus laevis) embryos: Biochemical dissection of control mechanisms.
Fruit fly (Drosophila melanogaster): Genetic analysis of growth and division coordination.
Cultured human cells: Molecular and microscopic studies.
Studying Cell-Cycle Progression
Cell-cycle stages can be determined through:
Microscopic observation of living cells.
Staining cells with DNA-binding dyes or antibodies.
Using visualizable molecules incorporated into newly synthesized DNA (e.g., EdU).
S-phase cells identified by EdU incorporation.
Estimating duration of S phase from labeled cells proportion.
Mitotic index (proportion of cells in mitosis) estimates M phase duration.
Fluorescently labeled proteins can track cell-cycle phases in living cells.
DNA content measurement via flow cytometry indicates cell stage (G1, S, G2/M).
Cell division
Begins with duplication of cell contents.
Followed by distribution of contents into two daugher cells.
Chromosome duplcation occurs in S phase, most other cell components duplicated continually throughout the cycle.
During M phase, replicated chromosomes are segregated into individual nuclei, cell then splits in two.
S and M phase separated by gap phases G1 and G2.
The Cell-Cycle Control System
The proteins of the control system are distinct from those involved in DNA replication and chromosome segregation.
The cell-cycle control system operates like a timer that triggers events in a set sequence.
Responds to information from the processes it controls; malfunctions during DNA synthesis delay progression to M phase.
Based on biochemical switches that initiate cell-cycle events.
Switches are binary (on/off) and trigger irreversible events.
Robust and reliable, adaptable. Can be modified to suit specific cell types or respond to specific intracellular or extracellular signals.
Major Regulatory Transitions:
Start (or Restriction Point): Commits to cycle entry and chromosome duplication.
G2/M Transition: Triggers early mitotic events, chromosome alignment.
Metaphase-to-Anaphase Transition: Stimulates sister chromatid separation.
Cyclin-Dependent Kinases (Cdks)
Central components of the cell-cycle control system.
Protein kinases whose activity rises and falls during the cycle, controlled by a complex array of proteins.
Cyclins are the most important Cdk regulators.
Cyclins undergo synthesis and degradation in each cell cycle. Cdk levels are generally constant.
Cyclical changes in cyclin levels cause cyclic assembly and activation of cyclin-Cdk complexes.
Three Major Classes of Cyclins:
G1/S-cyclins: Activate Cdks in late G1, trigger progression through Start, levels fall in S phase.
S-cyclins: Bind Cdks after Start, stimulate chromosome duplication, levels remain elevated until mitosis, control some early mitotic events.
M-cyclins: Activate Cdks that stimulate entry into mitosis at the G2/M transition, levels fall in mid-mitosis.
G1-cyclins govern G1/S-cyclin activities, controlling progression through Start.
Extracellular signals that stimulate cell proliferation act by increasing the production of G1-cyclins.
Yeast cells:
Single Cdk protein binds all cyclin classes, triggers different events by changing cyclin partners.
Vertebrate cells:
Four Cdks: Two interact with G1-cyclins, one with G1/S- and S-cyclins, one with S- and M-cyclins.
Cyclin binding is not enough for complete activation of Cdks.
Cdk-activating kinase (CAK) phosphorylates an amino acid near the Cdk active site.
Wee1 kinase inhibits Cdk activity by phosphorylation near the active site.
Cdc25 phosphatase increases Cdk activity by dephosphorylation.
Cdk inhibitor proteins (CKIs) govern G1/S- and S-Cdks early in the cell cycle.
Protein Phosphatases
They remove phosphate.
Protein phosphatase 2A (PP2A) is a critical regulator of Cdk substrates.
Consists of three subunits; activity is high during interphase (But Inhibited During Early Mitosis When M-Cdk Activity Rises).
When anaphase initiated M-.Cdk Declines: PP2A-B55 Reactivated to Promote Rapid Dephosphorylation of Cdk Substrates During Anaphase. and Telaphase.
Cdk Substrates
Cyclins direct Cdks to specific target proteins.
Single enzyme phosphorylates different targets in a specific order.
Ordering depends on the affinities of the interactions between the Cdk active site and the substrate.
Total amount of enzyme activity also important.
Positive Feedback
Cell-cycle control system generates switchlike, binary decisions through positive feedback.
The activation of M-Cdk at the G2/M transition:
M-cyclin accumulates during G2, leads to accumulation of M-Cdk complexes (primed and ready to act but suppressed).
Cdc25 removes inhibitory phosphates that restrain M-Cdk.
At the same time, inhibitory activity of kinase Weel is suppressed further ensuring increase of M-Cdk activity.
M-Cdk activities activates its own activator Cdc25 and inhibits the inhibitory kinase Weel - results in positive feedback.
The Anaphase-Promoting Complex/Cyclosome (APC/C)
Regulates the metaphase-to-anaphase transition.
It is a ubiquitin ligase enzyme family.
It can Polyubiquitylate specific target proteins - resulting in their destruction in proteasomes.
APC/C Catalyzes the Ubiquitylation and Destruction of Two Major Proteins:
First: Securin (Destruction in metaphase activates a protease that separates the sister-chromatid pairs).
Second: Destroying S- and M-cyclins (inactivates most Cdks in the cell).
APC/C (Activation)
Activity increases in mid-mitosis and remains high through G1
Activation depends on association with one of two activating subunits, Cdc20 or Cdh1.
Cdc20 - Acts first, in metaphase, to trigger the destruction of securin and cyclins resulting in chromosome segregation in anaphase.
Cdh1 - Maintains APC/C activity through late mitosis and G1.
SCF also employs another ubiquitin ligase.
The G1 Phase
A Stable State of Cdk Inactivity Is the Result
Resetting the cell-cycle control system, after Key Event in Plate M phase inactivation of Cdks the cell prepares for a new cell cycle.
During this phase, the cell grows and carries out its normal functions while also checking for DNA damage and ensuring all necessary resources are available for the subsequent phases.
This phase is crucial for maintaining genomic integrity and proper cellular function, as any errors detected may prompt the cell to pause and initiate repair mechanisms before progressing to the S phase.
This process is referred to as the G1 phase, where the cell also evaluates external signals, such as growth factors, that can influence its decision to divide.
In this phase, the cell not only assesses its internal conditions but also responds to external stimuli, which include physical space and nutrient availability. If conditions are favorable, the cell advances to the synthesis stage, or S phase, where DNA replication occurs. In contrast, if the cell detects unfavorable conditions or significant DNA damage, it may enter a resting state known as G0, where it remains metabolically active but does not prepare for division. This state allows the cell to conserve resources and avoid unnecessary division until conditions improve or damage is repaired, highlighting the cell's strategic approach to growth and division. At this point, the cell must undergo critical checkpoints to ensure that all preparations for division are adequate before entering the next phase, G2, where further growth occurs and final preparations are made for mitosis. During G2, the cell continues to grow and produces proteins necessary for mitosis, while also repairing any remaining damage to its DNA. This careful regulation ensures that genomic integrity is maintained and that the cell is fully equipped for the division process. Overall, the G2 phase is crucial for confirming that the cell has all the components needed for successful mitosis, emphasizing the importance of these checkpoints in preventing errors that could lead to malfunction or disease. Following G2, the cell enters the M phase, where the actual process of mitosis occurs, resulting in the division of the cell's nucleus, followed by cytokinesis, which divides the cytoplasm and completes the cell division cycle. In summary, the cell cycle is a tightly regulated sequence that ensures proper division and distribution of genetic material, with each phase playing a vital role in maintaining cellular health and function. The cell cycle is divided into several phases: G1, S, G2, and M, each characterized by specific events that contribute to cellular growth, DNA replication, and division.
G1 Phase: The first growth phase, where the cell grows in size and synthesizes proteins necessary for DNA replication, while also checking for any DNA damage. During this phase, the cell also produces organelles and increases its metabolic activity, setting the stage for the subsequent synthesis (S) phase where DNA replication occurs. S Phase: Characterized by the synthesis of DNA, where each chromosome is duplicated, resulting in two sister chromatids, which are essential for ensuring that each daughter cell receives an identical set of chromosomes. G2 Phase: The second growth phase, which involves further cell growth, the production of proteins for cell division, and the final preparations for mitosis, including the correction of any DNA replication errors.
M Phase: The mitotic phase where the cell undergoes division, encompassing both mitosis and cytokinesis, leading to the formation of two distinct daughter cells, each with an identical set of chromosomes.
Checkpoints: Critical control mechanisms are in place during the cell cycle to ensure proper progression and to prevent errors in cell division. These checkpoints monitor the cell's conditions and assess whether the processes of DNA replication and division are completed accurately, thereby safeguarding against cancerous growth and genetic defects.
Apoptosis: A programmed cell death mechanism that can be initiated if severe DNA damage is detected, preventing the proliferation of potentially harmful cells. This process serves as a safeguard to maintain cellular integrity and function, allowing the organism to eliminate damaged cells that may compromise overall health.
Interphase: The stage of the cell cycle where the cell prepares for division, consisting of three phases: G1 (gap 1), S (synthesis), and G2 (gap 2). During interphase, the cell grows, replicates its DNA, and produces the necessary proteins and organelles required for mitosis. Mitosis: The phase of the cell cycle where the actual cell division occurs, resulting in two daughter cells that are genetically identical to the original cell. Mitosis is further divided into four stages: prophase, metaphase, anaphase, and telophase, each characterized by specific changes in the cell's structure and chromosomal organization. Cytokinesis: The final stage of the cell cycle, where the cytoplasm divides, leading to the physical separation of the two daughter cells, completing the cell division process. Post-mitotic phase: The period following cytokinesis, where the daughter cells enter their respective interphase, preparing for their own cell cycle, and can differentiate into specific cell types based on the organism's needs. In summary, the cell cycle is a highly regulated process that ensures proper cell growth, DNA replication, and division, which are critical for maintaining healthy tissue function and organismal development. In addition to these phases, the cell cycle is also influenced by various checkpoints that monitor the integrity of the cell's DNA and overall health, preventing the division of damaged or unprepared cells. The checkpoints include the G1 checkpoint, which assesses cell size and DNA damage; the G2 checkpoint, which ensures DNA replication has been completed accurately; and the M checkpoint, which verifies proper chromosome alignment before mitosis proceeds. The regulation of these checkpoints is crucial, as they prevent the propagation of mutations and maintain genomic stability, thereby playing a significant role in cancer prevention and overall cellular health. Furthermore, disruptions in these checkpoints can lead to uncontrolled cell division, a hallmark of cancer, underscoring the importance of understanding the mechanisms that govern the cell cycle. Moreover, research into the specific proteins and enzymes involved in checkpoint regulation has provided insights into potential therapeutic targets for cancer treatment, highlighting the necessity of continued exploration in this field.
M Phase: The mitotic phase where the cell undergoes division, encompassing both mitosis and cytokinesis, leading to the formation of two distinct daughter cells, each with an identical set of chromosomes.