Cell Cycle and Mitosis

The Cell Cycle — An Overview

Part 1 of 7

Every living organism depends on cell division for growth, repair, and reproduction. The cell cycle is the ordered sequence of events that a cell undergoes from one division to the next.

A typical mammalian cell cycle lasts about 24 hours, though this varies enormously — some embryonic cells divide in 8 minutes, while liver cells may go years between divisions.

Phases of the Cell Cycle

The cell cycle consists of two major periods:

1. Interphase (~90% of the cell cycle)

• G$_1$ phase (Gap 1) — cell growth, organelle duplication, preparation for DNA synthesis
• S phase (Synthesis) — DNA replication; each chromosome is duplicated into two sister chromatids joined at the centromere
• G$_2$ phase (Gap 2) — continued growth, preparation for mitosis; error-checking of replicated DNA

2. Mitotic (M) phase (~10% of the cell cycle)

• Mitosis — division of the nucleus (karyokinesis) into two genetically identical daughter nuclei
• Cytokinesis — division of the cytoplasm to form two separate daughter cells

G$_0$ Phase: Some cells exit the cell cycle and enter a quiescent state called G$_0$. These cells are metabolically active but do not divide. Examples include neurons and mature muscle cells. Some G$_0$ cells can re-enter the cycle if stimulated (e.g., hepatocytes after liver damage).

Checkpoint — Cell Cycle Overview

Chromosome Organization at Each Stage

Understanding chromosome structure throughout the cell cycle is critical:

Before S phase (G$_1$):

• Each chromosome = 1 DNA double helix + histones = unreplicated chromosome
• Human cell: 46 unreplicated chromosomes

After S phase (G$_2$ and early mitosis):

• Each chromosome = 2 identical copies (sister chromatids) joined at the centromere = replicated chromosome
• Human cell: 46 replicated chromosomes (= 92 chromatids)
• The chromosome number does NOT change after replication — the cell still has 46 chromosomes

After mitosis:

• Sister chromatids separate → each becomes an independent chromosome
• Each daughter cell: 46 unreplicated chromosomes (identical to the original)

Key Distinction: "Chromosome number" counts centromeres, not chromatids. A replicated chromosome with two sister chromatids counts as ONE chromosome.

DNA packaging hierarchy:

1. DNA double helix (2 nm diameter)
2. DNA wraps around histone octamers → nucleosomes ("beads on a string," 11 nm)
3. Nucleosomes coil into 30 nm fiber (solenoid)
4. Looped domains of 30 nm fiber attached to a protein scaffold
5. Maximum condensation during metaphase → visible chromosomes (~1400 nm)

Checkpoint — Chromosome Counting

Key Terms — Cell Cycle

Match the Phase

Exit Ticket — Cell Cycle Phases

Interphase — Preparation for Division

Part 2 of 7

Interphase is often called the "resting phase," but this is a misnomer — the cell is extremely active during interphase. It is growing, producing proteins, duplicating organelles, and (during S phase) replicating its entire genome.

G$_1$ Phase — Growth and Preparation

G$_1$ is typically the longest and most variable phase of interphase:

Key events:

• Cell growth — increase in cell size and mass
• Synthesis of proteins, lipids, and carbohydrates
• Duplication of organelles (ribosomes, mitochondria, ER)
• Centriole duplication begins (in animal cells)
• Gene expression patterns establish the functional identity of the cell

G$_1$/S Checkpoint (Restriction Point):

• The most important checkpoint in the cell cycle
• The cell "decides" whether to commit to division
• Checks: adequate cell size, sufficient nutrients, growth factor signals, and intact DNA
• If the cell passes this checkpoint, it is committed to S phase and division
• If conditions are unfavorable, the cell enters G$_0$

Growth Factors: External signals (like PDGF, EGF, and insulin-like growth factor) bind to receptors and activate signaling cascades that promote passage through the G$_1$/S checkpoint. Cancer cells often have mutations that make them independent of growth factor signaling.

S Phase — DNA Replication

During S phase, the entire genome is copied:

Key events:

• Each chromosome is replicated by DNA polymerase using semi-conservative replication
• Replication begins at many origins of replication simultaneously (humans have ~30,000-50,000 origins)
• New histone proteins are synthesized and assembled onto the replicated DNA
• After replication, each chromosome consists of two sister chromatids joined at the centromere by cohesin proteins
• The centrosome (containing two centrioles in animal cells) is also duplicated

Replication timing:

• Not all chromosomes replicate simultaneously
• Euchromatin (active genes) replicates early in S phase
• Heterochromatin (condensed, inactive regions) replicates late in S phase
• Each origin of replication fires only ONCE per S phase (licensing system prevents re-replication)

DNA Content Notation: A cell in G$_1$ has 2n chromosomes and 2C DNA content. After S phase (in G$_2$), it has 2n chromosomes but 4C DNA content (each chromosome has doubled its DNA).

G$_2$ Phase — Final Preparation

G$_2$ is the final preparation period before mitosis:

Key events:

• Continued cell growth and protein synthesis
• Synthesis of proteins needed for mitosis (e.g., tubulin for the mitotic spindle)
• Final duplication of centrosomes completed
• G$_2$/M Checkpoint — the cell verifies:
  • All DNA has been completely and accurately replicated
  • DNA damage has been repaired
  • The cell is large enough to divide
  • Critical mitotic proteins are present

If DNA damage is detected: The checkpoint kinase ATM/ATR activates p53, which can halt the cell cycle (allowing time for repair) or trigger apoptosis (programmed cell death) if the damage is too severe to repair.

Checkpoint — Interphase

Key Terms — Interphase

Exit Ticket — Interphase

Mitosis — Dividing the Nucleus

Part 3 of 7

Mitosis is the division of the nucleus to produce two genetically identical daughter nuclei. It is a continuous process but is conventionally divided into four (or five) stages: prophase, prometaphase, metaphase, anaphase, and telophase.

Prophase

Key events:

• Chromatin condenses into visible chromosomes (each consisting of two sister chromatids joined at the centromere)
• Condensation is driven by condensin proteins that coil and compact the chromatin
• The mitotic spindle begins to form:
  • In animal cells: centrosomes (each with two centrioles) migrate toward opposite poles; asters (radial arrays of microtubules) form around them
  • In plant cells: spindle forms without centrioles (acentrosomal spindle)
• Nucleolus disappears (ribosomal RNA synthesis ceases)

Prometaphase

Key events:

• Nuclear envelope breaks down (fragments into vesicles)
• Spindle microtubules now access the chromosomes
• Kinetochores form at the centromere of each sister chromatid — these are protein complexes that serve as attachment points for spindle microtubules
• Kinetochore microtubules from opposite poles attach to the kinetochores of sister chromatids
• Chromosomes are moved by motor proteins along microtubules in a "search and capture" process

Metaphase

Key events:

• All chromosomes align at the metaphase plate (the equator of the cell, equidistant from both poles)
• Each chromosome is attached to kinetochore microtubules from BOTH poles (bipolar attachment)
• Spindle Assembly Checkpoint (SAC) — verifies that every kinetochore is properly attached to spindle microtubules from both poles before allowing anaphase to proceed

The Spindle Assembly Checkpoint is critical: If even one kinetochore is unattached, the checkpoint protein Mad2 inhibits the Anaphase-Promoting Complex (APC/C), preventing the cell from entering anaphase. Failure of this checkpoint leads to aneuploidy (abnormal chromosome number).

Anaphase

The shortest phase of mitosis, but the most dramatic:

Key events:

• Cohesin proteins holding sister chromatids together are cleaved by the enzyme separase
• Separase is activated when APC/C degrades securin (the inhibitor of separase)
• Sister chromatids separate and move toward opposite poles:
  • Anaphase A: Kinetochore microtubules shorten (depolymerize at the kinetochore end), pulling chromatids poleward
  • Anaphase B: Polar microtubules elongate, pushing the poles apart; motor proteins (dynein) pull on astral microtubules, moving poles apart
• Once separated, each chromatid is now called an independent chromosome

Telophase

Key events:

• Chromosomes arrive at opposite poles and begin to decondense (uncoil back into chromatin)
• Nuclear envelope re-forms around each set of chromosomes (from vesicles and ER membrane)
• Nucleolus reappears
• Spindle microtubules depolymerize
• The cell now contains two nuclei, each with a complete set of chromosomes

Summary of chromosome movement by microtubule type:

Checkpoint — Mitosis Stages

Key Terms — Mitosis

Match the Mitosis Stage

Exit Ticket — Mitosis

Cytokinesis — Dividing the Cytoplasm

Part 4 of 7

Cytokinesis is the division of the cytoplasm to produce two separate daughter cells. It typically begins during anaphase or telophase and overlaps with the final stages of mitosis.

The mechanism differs between animal and plant cells.

Cytokinesis in Animal Cells — Cleavage Furrow

Animal cells divide by cleavage:

1. A contractile ring of actin microfilaments and myosin II motor proteins assembles just beneath the plasma membrane at the former metaphase plate
2. The position of the contractile ring is determined by signals from the mitotic spindle (specifically, the central spindle — overlapping polar microtubules between the separating chromosomes)
3. Myosin II hydrolyzes ATP and slides along actin filaments, constricting the ring
4. This creates an inward indentation called the cleavage furrow
5. The furrow deepens progressively until the cell is pinched in two
6. Final separation (abscission) involves membrane fusion at the narrow bridge connecting the two cells

Why the middle? The position of the contractile ring is specified by signals from the spindle midzone and astral microtubules. The RhoA GTPase pathway activates myosin II and actin assembly at the equator. This ensures the cell divides between the two sets of chromosomes.

Cytokinesis in Plant Cells — Cell Plate

Plant cells cannot form a cleavage furrow because of their rigid cell wall. Instead, they build a new cell wall from the inside out:

1. Golgi-derived vesicles carrying cell wall materials (polysaccharides, glycoproteins) are transported along remaining spindle microtubules to the center of the cell
2. Vesicles fuse to form the cell plate, which grows outward from the center toward the existing cell wall
3. The cell plate matures into a new middle lamella (shared layer between adjacent cell walls) and regions of new cell wall (primary wall)
4. New plasma membrane lines each side of the cell plate
5. The cell plate eventually fuses with the existing cell wall, completing the division

Plasmodesmata (channels connecting adjacent plant cells) are established during cell plate formation — portions of ER become trapped in the forming plate, creating cytoplasmic bridges between daughter cells.

Checkpoint — Cytokinesis

Binary Fission in Prokaryotes

Prokaryotes (bacteria and archaea) do not undergo mitosis. They reproduce by binary fission:

1. The single circular chromosome is replicated starting from the origin of replication (oriC)
2. The two copies of the chromosome are moved to opposite ends of the cell (by proteins attached to the membrane, including ParA/ParB system)
3. The cell elongates
4. A septum (new cell wall and membrane) forms at the midpoint, guided by the FtsZ protein ring (a tubulin-like protein that forms a contractile ring analogous to the animal cell contractile ring)
5. The cell divides into two identical daughter cells

Key differences from mitosis:

Key Terms — Cytokinesis

Exit Ticket — Cytokinesis

Cell Cycle Regulation — Checkpoints and Cancer

Part 5 of 7

The cell cycle is tightly regulated to ensure accurate DNA replication and equal chromosome distribution. The control system relies on cyclins, cyclin-dependent kinases (Cdks), checkpoints, and tumor suppressors.

Loss of cell cycle control is the fundamental basis of cancer.

Cyclins and Cdks — The Engine of the Cell Cycle

Cyclin-dependent kinases (Cdks) are enzymes that phosphorylate target proteins to drive the cell through each phase. Cdks are only active when bound to a cyclin partner.

Key Cdk-cyclin complexes:

How cyclin-Cdk regulation works:

1. Cyclins are synthesized and degraded in a cyclical pattern — each cyclin accumulates during a specific phase
2. When cyclin levels are high, Cdks are activated
3. Activated Cdk phosphorylates target proteins, driving the next cell cycle event
4. After the event, cyclins are tagged for destruction by ubiquitin ligases (e.g., APC/C, SCF) and degraded by the proteasome
5. Cdk activity drops, resetting the system

MPF (Maturation/Mitosis Promoting Factor): The Cyclin B-Cdk1 complex was originally discovered in frog oocytes as the factor that triggers entry into M phase. Its discovery by Leland Hartwell, Tim Hunt, and Paul Nurse earned the 2001 Nobel Prize in Physiology or Medicine.

Checkpoint Summary

The Rb Pathway (G$_1$/S control)

1. Rb (Retinoblastoma protein) normally INHIBITS E2F transcription factors
2. When E2F is inhibited, genes needed for S phase entry are NOT transcribed
3. Growth factor signaling activates Cyclin D-Cdk4/6, which phosphorylates Rb
4. Phosphorylated Rb releases E2F
5. E2F activates transcription of genes for DNA replication (DNA polymerase, thymidine kinase, Cyclin E, etc.)
6. Cyclin E-Cdk2 further phosphorylates Rb, creating a positive feedback loop that commits the cell to S phase

Checkpoint — Regulation

Cancer — Loss of Cell Cycle Control

Cancer results from the accumulation of mutations in genes that regulate the cell cycle. Two categories of genes are involved:

1. Proto-oncogenes → Oncogenes (gain-of-function mutations)

• Proto-oncogenes are normal genes that PROMOTE cell division (e.g., growth factors, growth factor receptors, signal transduction proteins, cyclins)
• A mutation that makes the gene product hyperactive or overexpressed converts it into an oncogene
• Only ONE mutant allele is needed (dominant)
• Examples:
  • Ras — GTPase in growth factor signaling; mutant Ras is stuck "on" (~30% of all cancers)
  • HER2 — growth factor receptor overexpressed in some breast cancers
  • Myc — transcription factor that drives cell proliferation

2. Tumor suppressor genes (loss-of-function mutations)

• Normal products INHIBIT cell division, promote apoptosis, or repair DNA
• BOTH alleles must be inactivated (recessive — Knudson two-hit hypothesis)
• Examples:
  • p53 — "guardian of the genome"; halts the cycle for DNA repair or triggers apoptosis; mutated in >50% of cancers
  • Rb — blocks E2F-mediated S phase entry; loss causes retinoblastoma
  • BRCA1/BRCA2 — DNA repair (homologous recombination); mutations increase breast/ovarian cancer risk

Hallmarks of Cancer (Hanahan & Weinberg): sustained proliferative signaling, evading growth suppressors, resisting cell death, enabling replicative immortality (telomerase), inducing angiogenesis, activating invasion and metastasis.

Checkpoint — Cancer

Key Terms — Regulation and Cancer

Exit Ticket — Regulation

Problem-Solving Workshop — Cell Cycle

Part 6 of 7

This workshop applies cell cycle and mitosis concepts to experimental scenarios commonly seen on the AP Biology exam.

Scenario 1: Mitotic Index Calculation

A student observes 200 onion root tip cells under a microscope and counts the number of cells in each stage:

Mitotic Index = (cells in mitosis / total cells) $\times$ 100

$\text{Mitotic Index} = \frac{14 + 6 + 4 + 6}{200} \times 100 = \frac{30}{200} \times 100 = 15\%$

The mitotic index tells us the proportion of cells actively dividing. A higher mitotic index indicates more rapid cell proliferation.

The relative time in each phase is proportional to the number of cells observed:

• If the total cell cycle is 24 hours, and 15% of cells are in mitosis, then mitosis takes ~3.6 hours
• Prophase (14/200 = 7%) $\approx$ 1.68 hours
• Anaphase (4/200 = 2%) $\approx$ 0.48 hours (shortest phase — confirmed by observation)

Scenario 1 Questions

Scenario 2: Cell Fusion Experiment

Researchers fuse cells in different phases and observe the results:

Experiment A: S-phase cell fused with G$_1$-phase cell

• Result: The G$_1$ nucleus immediately enters S phase (begins DNA replication)
• Interpretation: S-phase cells contain factors (Cyclin E/A-Cdk2) that can drive a G$_1$ nucleus into S phase

Experiment B: M-phase cell fused with G$_1$-phase cell

• Result: The G$_1$ nucleus undergoes premature chromosome condensation (PCC) — but the chromosomes are single-chromatid (unreplicated), leading to chromosome damage
• Interpretation: M-phase cells contain MPF (Cyclin B-Cdk1) that forces any nucleus to enter M phase regardless of DNA replication status

Experiment C: M-phase cell fused with S-phase cell

• Result: The S-phase chromosomes show premature condensation of partially replicated DNA — pulverized chromosomes (chromosome fragmentation)
• Interpretation: Forcing M phase entry on incompletely replicated DNA is catastrophic

These experiments demonstrated that cell cycle progression is controlled by diffusible cytoplasmic factors (cyclins and Cdks), not by fixed programs in the nucleus alone.

Scenario 2 Questions

Apply Your Knowledge

Exit Ticket — Workshop

AP Review — Cell Cycle and Mitosis

Part 7 of 7

Comprehensive AP-exam-style questions integrating all cell cycle and mitosis concepts.

Key Principles Summary

1. The cell cycle consists of interphase (G$_1$, S, G$_2$) and M phase (mitosis + cytokinesis)
2. Mitosis produces two genetically identical daughter cells (preserving chromosome number)
3. Cyclin-Cdk complexes drive progression through each phase; cyclin degradation resets the system
4. Checkpoints (G$_1$/S, G$_2$/M, SAC) ensure accuracy before committing to the next phase
5. Cancer results from mutations in proto-oncogenes (gain of function) and tumor suppressors (loss of function)
6. Cytokinesis uses a cleavage furrow (animal) or cell plate (plant)

AP-Style Questions — Set 1

AP-Style Questions — Set 2

Comprehensive Matching

Final Review

Final Exit Ticket