Replication and division of nuclei and cells

The cell cycle is the regular sequence of events between one cell division and the next. A typical mammalian cell completes one cycle in about 24 hours, of which mitosis and cytokinesis occupy less than an hour — the rest is interphase. The cell cycle has two main parts:

Interphase — the period of cell growth, DNA replication and preparation. Although chromosomes are not visible, the cell is metabolically very active. Interphase is subdivided into three phases:

• G1 (Gap 1). The cell grows in size, synthesises new proteins and organelles (more ribosomes, mitochondria, ER), and carries out normal metabolic functions. The first major checkpoint at the end of G1 (the 'restriction point') determines whether the cell will commit to a full division cycle, exit to G0 (a non-dividing resting state), or undergo differentiation.
• S (Synthesis) phase. DNA replication occurs. Each DNA molecule is copied semi-conservatively, so each chromosome ends S phase as two identical sister chromatids joined at a centromere. The amount of DNA per cell doubles. Histone proteins are also synthesised, so the chromatin can be packaged.
• G2 (Gap 2). Further growth and preparation for mitosis: synthesis of microtubule subunits for the spindle, replication of centrosomes (animal cells), and a DNA damage checkpoint that verifies S-phase replication was accurate.

M phase (mitosis) — nuclear division: chromosomes condense, line up on the equator, sister chromatids separate, and two genetically identical daughter nuclei form. Four stages: prophase, metaphase, anaphase, telophase.

Cytokinesis — division of the cytoplasm. Follows mitosis. Cleavage furrow in animal cells; cell plate in plant cells. Two daughter cells are produced.

After cytokinesis, the new daughter cells enter G1 and the cycle repeats — or they may exit to G0 and differentiate (most adult somatic cells, e.g. neurones, muscle fibres).

Mitosis produces two daughter nuclei that are genetically identical to each other and to the parent nucleus, each with the same chromosome number. This faithful copying is the basis of several essential biological roles.

Growth. Multicellular organisms grow from a single fertilised egg to an adult of many trillions of cells by repeated mitotic divisions. Because all somatic cells share the same genome, every cell carries the full set of instructions for the organism. Tissue-specific gene expression then allows specialisation while preserving genetic identity.

Repair and replacement. Damaged or worn-out cells are replaced by mitotic divisions of healthy neighbours:

• Skin — basal cells of the epidermis divide constantly to replace cells lost from the surface (turnover ~4 weeks).
• Gut lining — epithelial cells in the crypts of Lieberkuhn divide every 3-6 days.
• Bone marrow — haematopoietic stem cells produce $\sim 2 \times 10^{11}$ red blood cells per day.
• Wound healing — fibroblasts and epithelial cells divide to seal a cut.

Asexual reproduction. Many organisms produce genetically identical offspring (clones) by mitosis:

• Binary fission in Amoeba, Paramecium and other unicellular eukaryotes.
• Budding in Hydra and yeast.
• Vegetative propagation in plants — strawberry runners, potato tubers, onion bulbs, bamboo rhizomes.

This is useful in stable, favourable environments because a successful genotype is preserved exactly — there is no risk of producing inferior offspring from genetic recombination.

Maintains chromosome number. Because sister chromatids separate so each daughter cell receives one chromatid per chromosome, the diploid chromosome number is maintained across generations of somatic cells. This is essential for stable inheritance and the consistent functioning of cells.

Note. Mitosis does not generate genetic variation. Variation arises only through meiosis (which produces gametes with reshuffled and reduced chromosomes) and fertilisation.

The cell cycle is normally tightly regulated so that cells divide only when needed and stop dividing when conditions are unfavourable. Regulation is provided by:

• Proto-oncogenes — encode proteins that promote cell division (growth factors, growth-factor receptors, cyclins and cyclin-dependent kinases). They act as accelerators.
• Tumour suppressor genes — encode proteins that inhibit the cell cycle or trigger apoptosis (programmed cell death) when DNA is damaged. They act as brakes. The most famous is p53 ('the guardian of the genome'), mutated in over half of all human cancers; another is RB (the retinoblastoma gene).

Cancer arises from mutations in these genes:

• A mutation in a proto-oncogene can convert it into a permanently active oncogene that drives continuous division.
• A mutation that inactivates a tumour suppressor gene removes its brake on the cycle, so damaged cells are no longer arrested or eliminated.

Cancer is a multistep disease: typically several mutations must accumulate in the same cell lineage (usually at least one oncogene activation AND multiple tumour-suppressor inactivations) before cells fully escape normal control. Carcinogens (mutagens that cause cancer) accelerate the accumulation of these mutations. Major categories include:

• Ionising radiation — X-rays, gamma rays from radioactive decay (e.g. radon gas, Chernobyl fallout).
• UV radiation — sunlight UV-B causes skin cancers (basal cell carcinoma, squamous cell carcinoma, melanoma).
• Chemicals — many in tobacco smoke (benzopyrene), asbestos fibres, alcohol, certain dyes, aflatoxins from contaminated grains.
• Viruses — HPV (human papilloma virus) is responsible for the great majority of cervical cancer cases; EBV is linked to Burkitt lymphoma; Hepatitis B and C viruses are linked to liver cancer.

A tumour is a mass of cells produced by uncontrolled division. Tumours are classed as:

• Benign — stays localised, surrounded by a connective-tissue capsule, does not invade other tissues, does not metastasise. Often harmless (e.g. warts, lipomas) and removable by surgery.
• Malignant — invades surrounding tissues by secreting enzymes that break down extracellular matrix, and can shed cells that travel via blood or lymph to start secondary tumours elsewhere. This spread is called metastasis. Only malignant tumours are clinically described as cancer.

Treatment options include surgery, radiotherapy (ionising radiation targeting tumour cells), chemotherapy (drugs that kill rapidly dividing cells), targeted molecular therapies (e.g. monoclonal antibodies against tumour antigens) and immunotherapy.

Telomeres are repetitive DNA-protein structures at the ends of linear eukaryotic chromosomes. In vertebrates, the telomeric DNA sequence is the hexanucleotide repeat TTAGGG, repeated thousands of times. Specialised proteins (the 'shelterin' complex) bind this sequence to form a protective cap.

Functions of telomeres:

• Protect chromosome ends from being recognised as broken DNA (which would trigger repair, fusion or apoptosis).
• Prevent fusion of chromosome ends to one another.
• Allow complete replication of the coding part of the chromosome despite the 'end-replication problem' (DNA polymerase cannot replicate the very end of a linear DNA molecule, so some sequence is lost each round — but if the lost sequence is telomere, no coding information is lost).

Shortening and the Hayflick limit. Because of the end-replication problem, telomeres shorten by ~50-100 base pairs at every cell division. After many divisions, telomeres become too short, the protection fails, and the cell stops dividing — entering a non-dividing state called senescence. Leonard Hayflick (1961) discovered that normal human somatic cells in culture can undergo only ~40-60 divisions before reaching this 'Hayflick limit'. Telomere shortening therefore contributes to cellular ageing.

Telomerase. A ribonucleoprotein enzyme (with both RNA and protein components) that extends telomeres by adding TTAGGG repeats to the 3' end of the DNA. Telomerase is normally active only in:

• Germline cells (sperm, eggs) — telomeres must not shorten between generations.
• Stem cells — must divide many times throughout life.
• Many cancer cells — over 85% of cancers reactivate telomerase, giving them effectively unlimited replicative capacity. This is one of the changes that makes cancer cells different from normal cells, and telomerase inhibitors are an active area of cancer-drug research.

A stem cell is an undifferentiated cell that can:

1. Self-renew — divide by mitosis to produce more stem cells, AND
2. Differentiate — give rise to one or more specialised cell types.

Both criteria must be met. Stem cells are classified by their potency (the range of cell types they can produce):

Totipotent. Can give rise to every cell type, including the extra-embryonic tissues (placenta, umbilical cord). In humans, only the zygote and the cells of the very early embryo (up to about the 8-cell morula stage, ~3 days after fertilisation) are totipotent. A single totipotent cell can generate a complete organism.

Pluripotent. Can give rise to all the cell types of the body (the three germ layers — ectoderm, mesoderm, endoderm — and all their derivatives) but not the extra-embryonic tissues. Found in the inner cell mass of the blastocyst (~5 days after fertilisation). Embryonic stem cells (ESCs) used in research are pluripotent. Induced pluripotent stem cells (iPSCs) — adult cells genetically reprogrammed to a pluripotent state — also fall in this category.

Multipotent. Can give rise to a limited range of related cell types, usually within one tissue lineage. Most adult tissue-specific stem cells are multipotent. Important examples:

• Haematopoietic stem cells in red bone marrow give rise to all blood-cell types (erythrocytes, granulocytes, lymphocytes, platelets).
• Mesenchymal stem cells in connective tissue give rise to bone, cartilage and fat cells.
• Neural stem cells in the brain give rise to neurones and glia (limited).
• Epithelial stem cells in the basal layer of skin and the crypts of the gut continuously replace surface cells.

Importance of stem cells.

• Growth. Embryonic stem cells underpin all of development from zygote to adult.

• Repair and replacement. Adult stem cells maintain renewing tissues throughout life — skin, gut, blood, bone marrow, hair follicles.

• Medicine. Stem-cell therapies are already used clinically:

  • Bone marrow transplants for leukaemia — donor haematopoietic stem cells repopulate the patient's bone marrow.
  • Skin grafts grown from epidermal stem cells for burns patients.
  • Cornea repair from limbal stem cells.

  Active research areas include type-1 diabetes ($\beta$-cells from pluripotent stem cells), Parkinson's disease (dopaminergic neurones), spinal-cord injury, macular degeneration, and heart failure.

• Ethical considerations. Embryonic stem cells require destruction of human embryos, which is ethically controversial. iPSC technology (Yamanaka, 2006 — Nobel Prize 2012) addresses this by reprogramming adult cells (e.g. skin fibroblasts) back to a pluripotent state using four transcription factors (Oct4, Sox2, Klf4, c-Myc).

The cell cycle is regulated by checkpoints — surveillance mechanisms that halt the cycle if conditions are unfavourable or if errors are detected. Three main checkpoints operate:

G1 / restriction checkpoint. At the end of G1, the cell asks: is the cell large enough? Are there sufficient nutrients? Is the DNA undamaged? Are growth factors present? If yes, the cell commits to a full division cycle. If no, the cell can exit to G0 (resting state) or trigger apoptosis if damage is severe. p53 plays a central role here — it senses DNA damage and arrests the cycle to allow repair, or triggers apoptosis if repair fails.

G2 / DNA damage checkpoint. At the end of G2 (after S phase), the cell verifies that DNA replication is complete and accurate. If errors are detected, the cycle pauses to allow repair.

Metaphase / spindle assembly checkpoint. During metaphase, the cell verifies that every chromosome is correctly attached to spindle fibres from both poles before allowing anaphase to begin. This prevents unequal distribution of chromosomes (aneuploidy).

Cyclins and CDKs. Progress through the cycle is driven by waves of cyclin proteins that bind to cyclin-dependent kinases (CDKs). Each cyclin-CDK combination phosphorylates the proteins needed for the next phase. Cyclins are degraded at the end of their phase, returning the CDK to an inactive state.

Why this matters for cancer. Almost all cancers involve loss of one or more checkpoints — most often p53. Without functioning checkpoints, cells continue to divide despite DNA damage, accumulating further mutations and progressing towards malignancy.