Cell Biology BZO2T04

Syllabus topics (RTMNU)

• 1.1 Ultra-structure of Plasma membrane and it's chemical composition
• 1.2 Endomembrane system: Protein sorting, transport, exocytosis, endocytosis
• 1.3 Cell-cell interaction: Cell adhesion molecules, cellular junction
• 1.4 Golgi Complex: Structure and functions.

1.1 Ultra-structure of plasma membrane and its chemical composition

The plasma membrane (cell membrane, plasmalemma) is the thin, living, selectively permeable boundary of every cell. Under the electron microscope it looks like a trilaminar (three-layered) "railway track": two dark lines with a light line between them, about 7.5 nm (75 Å) thick.

Chemical composition

• Phospholipids are amphipathic: a hydrophilic (water-loving) phosphate head and two hydrophobic fatty-acid tails. In water they form a bilayer, heads outside, tails inside.
• Cholesterol fits between phospholipids and controls fluidity (makes the membrane less fluid at high temperature and prevents freezing at low temperature).
• Glycocalyx (cell coat) on the outer surface helps in cell recognition, blood groups and protection.

Models of membrane structure

1. Lipid bilayer – Gorter and Grendel (1925): lipid from RBC membranes covered twice the cell surface area, so the membrane is a bilayer.
2. Sandwich model – Danielli and Davson (1935): lipid bilayer sandwiched between two continuous layers of protein (protein–lipid–protein).
3. Unit membrane model – J. D. Robertson (1959): all biological membranes have the same trilaminar structure (dark–light–dark) seen in EM.
4. Fluid mosaic model – S. J. Singer and G. L. Nicolson (1972): the accepted model.

Fluid mosaic model – key points

• The lipid bilayer is quasi-fluid; lipids move sideways (lateral diffusion) quickly, but flip-flop from one layer to the other is rare.
• Proteins float in the lipid "sea" like icebergs, forming a mosaic.
• Peripheral (extrinsic) proteins: loosely attached on the surface; easily removed.
• Integral (intrinsic) proteins: embedded partly or fully; those that span the whole bilayer are transmembrane proteins (e.g. channels, carriers).
• The membrane is asymmetric: the two faces differ in lipids, proteins and carbohydrate (carbohydrate only on the outer face).
• Evidence: Frye and Edidin (1970) fused human and mouse cells; their labelled membrane proteins mixed within ~40 minutes, proving fluidity.

Diagram (describe in exam): draw two rows of phospholipids (circles with two tails) facing each other; show an integral protein passing through both rows, a peripheral protein on the inner surface, cholesterol between tails and branched carbohydrate chains on the outer side; label all parts.

Functions: selective permeability, transport (diffusion, osmosis, active transport by pumps like Na⁺/K⁺-ATPase), cell recognition, receptor for hormones, cell adhesion, endocytosis and exocytosis.

1.2 Endomembrane system: protein sorting, transport, exocytosis, endocytosis

The endomembrane system is a group of membrane-bound organelles that work together and exchange material by vesicles: nuclear envelope, endoplasmic reticulum (ER), Golgi complex, lysosomes, vacuoles/endosomes and the plasma membrane. Mitochondria, chloroplasts and peroxisomes are not part of it because their functions are not coordinated with these.

Protein sorting (targeting)

• All proteins start on ribosomes in the cytosol. Their final address is decided by a signal sequence (short amino-acid "zip code"). This is the signal hypothesis of Günter Blobel (Nobel Prize 1999).
• Co-translational (secretory) pathway: signal peptide at N-terminus → recognised by Signal Recognition Particle (SRP) → translation pauses → SRP binds SRP receptor on rough ER → ribosome docks on translocon → growing chain enters ER lumen → signal peptidase removes signal. Proteins for ER, Golgi, lysosome, plasma membrane and secretion follow this route.
• Post-translational pathway: proteins made on free ribosomes go to nucleus (nuclear localisation signal), mitochondria, peroxisomes (signal SKL at C-terminus) or stay in cytosol.
• Retention signal KDEL keeps soluble ER proteins in the ER. Mannose-6-phosphate tag sends acid hydrolases to lysosomes.

Vesicular transport

Route of a secretory protein: RER → transition vesicles → cis Golgi → medial → trans Golgi → secretory vesicle → plasma membrane (G. Palade's pulse-chase experiment on pancreatic cells proved this route).

Vesicles fuse with the correct target using SNARE proteins (v-SNARE on vesicle pairs with t-SNARE on target) and Rab GTPases.

Exocytosis

• Fusion of a vesicle with the plasma membrane releasing its contents outside; it also adds membrane to the cell surface.
• Constitutive – continuous, unregulated (e.g. collagen, mucus, membrane proteins).
• Regulated – stored in granules, released on a signal, usually a rise in Ca²⁺ (e.g. insulin from β-cells, neurotransmitters, digestive enzymes).

Endocytosis

• Intake of material by in-folding of the plasma membrane to form a vesicle.
• Phagocytosis ("cell eating"): large solid particles, bacteria; forms phagosome (e.g. macrophages, neutrophils, Amoeba).
• Pinocytosis ("cell drinking"): fluid and dissolved solutes in small vesicles.
• Receptor-mediated endocytosis: specific molecules bind receptors in clathrin-coated pits (e.g. LDL-cholesterol uptake; defect causes familial hypercholesterolaemia).
• Endocytic vesicle → early endosome → late endosome → lysosome (digestion).

1.3 Cell–cell interaction: cell adhesion molecules, cellular junctions

Cell adhesion molecules (CAMs)

Cellular junctions (in animal cells)

1. Tight junction (zonula occludens) – membranes of neighbouring cells fused by proteins claudin and occludin; seals the gap, stops leakage between cells. Example: intestinal epithelium, blood–brain barrier.
2. Adherens junction (zonula adherens) – belt of cadherins linked inside to actin filaments.
3. Desmosome (macula adherens) – button-like spot weld; cadherins (desmoglein, desmocollin) linked to intermediate filaments (keratin) through dense plaques. Gives mechanical strength; abundant in skin and heart muscle.
4. Hemidesmosome – half desmosome joining cell to basal lamina via integrins.
5. Gap junction (nexus) – channels made of six connexin proteins (a connexon); allow ions and small molecules (<1000 Da) to pass directly; electrical coupling in heart and smooth muscle.

Junctional complex in intestinal epithelium = tight junction + adherens junction + desmosome, from top to bottom. (Plant cells communicate through plasmodesmata.)

1.4 Golgi complex: structure and functions

Discovered by Camillo Golgi (1898) in nerve cells of barn owl by silver (metallic impregnation) staining. Also called dictyosome (in plants), Golgi apparatus, lipochondria.

Structure (EM)

• Cisternae: 4–8 flattened, curved, membrane sacs stacked in parallel, with swollen margins.
• Tubules: network arising from cisternal margins.
• Vesicles: small transport vesicles (~40–80 nm) and large secretory vacuoles.
• Polarity: the convex cis (forming) face faces the ER and receives vesicles; the concave trans (maturing) face faces the plasma membrane and gives off vesicles. Middle is the medial region. TGN (trans-Golgi network) is the sorting station.

Functions

• Glycosylation: completes modification of N-linked sugars and adds O-linked sugars → glycoproteins.
• Sorting and packaging of proteins into vesicles for secretion, lysosomes or membrane.
• Formation of lysosomes (primary lysosomes bud from the trans face).
• Secretion – zymogen granules, hormones, mucus.
• Formation of acrosome of sperm during spermiogenesis.
• Synthesis of glycolipids, sulphation of proteins, formation of yolk; in plants, cell plate and cell wall material.

Likely exam questions

Q1. Describe the fluid mosaic model of plasma membrane with a labelled diagram. (Long)

• Proposed by Singer and Nicolson (1972); earlier models: Danielli–Davson, Robertson's unit membrane.
• Quasi-fluid lipid bilayer; integral, transmembrane and peripheral proteins form a mosaic.
• Cholesterol controls fluidity; glycocalyx on outer face; membrane is asymmetric.
• Evidence: Frye–Edidin cell fusion experiment. Diagram with labels; functions.

Q2. Explain the structure of Golgi complex and add a note on its functions. (Long)

• Camillo Golgi, 1898; cisternae, tubules, vesicles, vacuoles.
• Cis (forming) face towards ER, trans (maturing) face and TGN towards plasma membrane.
• Functions: glycosylation, sorting and packaging, secretion, lysosome and acrosome formation, cell plate in plants.

Q3. What is receptor-mediated endocytosis? (Short)

Uptake of specific molecules (e.g. LDL) that bind receptors gathered in clathrin-coated pits; the pit pinches off as a coated vesicle, loses its coat and fuses with an endosome.

Q4. Write a note on desmosome. (Short)

A button-like anchoring junction (macula adherens): cadherins (desmoglein, desmocollin) join neighbouring membranes, and dense plaques inside link to keratin intermediate filaments. Gives strength to skin and heart muscle.

Q5. What is the signal hypothesis? (Short)

Günter Blobel's idea that a signal peptide at the N-terminus of a new protein is recognised by SRP, which takes the ribosome to the RER so the protein enters the ER lumen; signal peptidase then removes the signal.

Exam tip

Draw the fluid mosaic diagram neatly with at least six labels; the same diagram scores in questions on membrane structure, transport and composition.

Syllabus topics (RTMNU)

• 2.1 Cytoskeleton: Microtubules, Microfilaments, intermediate filaments.
• 2.2 Mitochondria: Structure and Oxidative phosphorylation, ETS.
• 2.3 Peroxisome: Structure and functions.
• 2.4 Ribosome: Structure, types, Lake's model and functions.

2.1 Cytoskeleton: microtubules, microfilaments, intermediate filaments

The cytoskeleton is a network of protein filaments in the cytoplasm of eukaryotic cells. It gives shape and support, anchors organelles, and helps in movement of the cell and of material inside it. It is dynamic: filaments are built and broken constantly.

Microtubules – key points

• Grow from the MTOC (microtubule organising centre) – the centrosome, with γ-tubulin rings; minus end at MTOC, plus end grows outward.
• Dynamic instability: rapid growth and shrinkage at the plus end.
• Centrioles and basal bodies: 9 triplets (9 + 0). Cilia and flagella axoneme: 9 + 2 arrangement of doublets with dynein arms.
• Drug colchicine blocks microtubule assembly (arrests cells at metaphase – used in karyotyping); taxol stabilises them.

Microfilaments – key points

• Globular G-actin polymerises into filamentous F-actin.
• Actin + myosin = sliding of filaments in muscle; contractile ring in cytokinesis; core of microvilli.
• Cytochalasin blocks actin polymerisation.

Intermediate filaments – key points

• Most stable, tough, tension-bearing; tissue-specific (keratin in epithelium, desmin in muscle, neurofilaments in neurons, vimentin in fibroblasts, lamins in nucleus of all cells).

2.2 Mitochondria: structure, oxidative phosphorylation, ETS

Mitochondria are the "power house of the cell" – site of aerobic respiration and ATP production. First seen by Kölliker (1857); named mitochondrion by C. Benda (1898); stained by Janus green B. Number: one to thousands per cell (more in active cells like muscle, liver). Absent in mature RBCs of mammals.

Structure

• Rod or sausage shaped, 0.5–1 µm wide, 1–10 µm long; double membrane.
• Outer membrane: smooth, freely permeable due to porin channels.
• Inner membrane: folded into cristae; impermeable to most ions; rich in protein; contains cardiolipin; carries the electron transport chain and F₀–F₁ particles (oxysomes / elementary particles of Fernández-Morán) – ATP synthase.
• Intermembrane space (outer chamber) between the two membranes.
• Matrix (inner chamber): enzymes of Krebs cycle and fatty-acid oxidation, circular double-stranded DNA, 70S ribosomes, RNA.
• Because of own DNA and ribosomes, mitochondria are semi-autonomous; endosymbiotic theory says they came from aerobic bacteria.

Electron transport system (ETS) – inner membrane

1. Complex I – NADH dehydrogenase (NADH-CoQ reductase): takes 2e⁻ from NADH (FMN, Fe–S); pumps 4 H⁺.
2. Complex II – succinate dehydrogenase: takes electrons from FADH₂ (succinate); pumps no H⁺.
3. Ubiquinone (coenzyme Q): mobile carrier in the lipid layer.
4. Complex III – cytochrome bc₁ complex: passes electrons to cytochrome c; pumps 4 H⁺.
5. Cytochrome c: small mobile protein on outer face of inner membrane.
6. Complex IV – cytochrome c oxidase (cyt a, a₃, Cu): gives electrons to O₂, the final electron acceptor, forming water (½O₂ + 2H⁺ + 2e⁻ → H₂O); pumps 2 H⁺.

Flow: NADH → I → CoQ → III → cyt c → IV → O₂. FADH₂ enters at II → CoQ.

Oxidative phosphorylation – chemiosmotic theory

• Proposed by Peter Mitchell (1961), Nobel Prize 1978.
• Energy from electron flow pumps H⁺ from matrix into intermembrane space → proton gradient (proton motive force).
• H⁺ flow back into matrix through the F₀ channel of ATP synthase; the F₁ head makes ATP: ADP + Pi → ATP.
• Yield: about 3 ATP per NADH and 2 ATP per FADH₂ (classical values; modern values 2.5 and 1.5). Total from one glucose: 38 ATP (classical) / ~30–32 (modern).
• Inhibitors: rotenone (I), antimycin A (III), cyanide and carbon monoxide (IV), oligomycin (ATP synthase). Uncouplers like 2,4-dinitrophenol and thermogenin in brown fat allow electron flow without ATP – energy lost as heat.

2.3 Peroxisome: structure and functions

• Discovered by J. Rhodin (1954) as "microbodies"; named peroxisome by Christian de Duve (1965).
• Small (0.5–1 µm), round, bounded by a single membrane; granular matrix, often a dense crystalline core (nucleoid) of urate oxidase in many animals (absent in humans).
• No DNA or ribosomes; proteins imported from cytosol (signal SKL). Multiply by division.
• Abundant in liver and kidney cells.

Functions

• Oxidases produce hydrogen peroxide: RH₂ + O₂ → R + H₂O₂.
• Catalase (marker enzyme) breaks it: 2H₂O₂ → 2H₂O + O₂; also detoxifies alcohol, phenol and formaldehyde (peroxidative reaction: H₂O₂ + R′H₂ → R′ + 2H₂O).
• β-oxidation of very long-chain fatty acids.
• Synthesis of plasmalogens (myelin lipids) and bile acids.
• Purine breakdown (uric acid → allantoin in many animals).
• In plants: photorespiration (leaf peroxisomes) and glyoxysomes (glyoxylate cycle, fat → sugar in seeds).
• Disorder: Zellweger syndrome (peroxisomes fail to form).

2.4 Ribosome: structure, types, Lake's model and functions

Ribosomes are small, non-membranous ribonucleoprotein particles – the site of protein synthesis. First observed by Claude (1943) as microsomes; described by George Palade (1955) ("Palade granules").

Structure and composition

• Made of rRNA (~60%) and protein (~40%); two unequal subunits – large and small – which join only during protein synthesis. Mg²⁺ (~0.001 M) is needed to hold the subunits together.
• Many ribosomes on one mRNA = polysome (polyribosome).
• Found free in cytoplasm, attached to RER, and inside mitochondria and chloroplasts.

Types

S = Svedberg unit (sedimentation coefficient in an ultracentrifuge); S values are not additive, so 50S + 30S = 70S.

Lake's model (James A. Lake, 1976)

• A three-dimensional model of the E. coli 70S ribosome made from electron micrographs.
• Small (30S) subunit: elongated, like an animal with a head, base and a platform; a cleft separates head and platform – mRNA binds here.
• Large (50S) subunit: like an armchair: a rounded body with three projections – a central protuberance in the middle, a ridge on one side and a stalk on the other; the peptidyl transferase site lies near the central protuberance.
• The two subunits fit so that a tunnel is left between them for mRNA, and an exit tunnel runs through the large subunit for the growing polypeptide.
• Sites: A site (aminoacyl-tRNA), P site (peptidyl-tRNA), E site (exit).

Functions

• Translation of mRNA into protein; the large subunit's 23S rRNA acts as peptidyl transferase (a ribozyme) forming peptide bonds.
• Free ribosomes make cytosolic proteins; RER-bound ribosomes make secretory, membrane and lysosomal proteins.

Likely exam questions

Q1. Describe the ultrastructure of mitochondria and explain oxidative phosphorylation. (Long)

• Double membrane, porins, cristae, F₀–F₁ particles, intermembrane space, matrix with circular DNA and 70S ribosomes.
• ETS: complexes I–IV, CoQ, cytochrome c, O₂ as final acceptor.
• Mitchell's chemiosmotic theory: proton gradient drives ATP synthase; ATP yield; inhibitors and uncouplers.

Q2. Compare microtubules, microfilaments and intermediate filaments. (Long)

• Diameter 25, 7 and 10 nm; proteins tubulin, actin, keratin/vimentin/lamins.
• Polarity, GTP/ATP use, motor proteins (kinesin, dynein, myosin).
• Functions: spindle and cilia; muscle contraction and microvilli; mechanical strength and nuclear lamina.

Q3. Describe Lake's model of ribosome. (Short)

J. A. Lake (1976) 3-D model of the E. coli 70S ribosome: 30S subunit with head, base and platform; 50S subunit with central protuberance, ridge and stalk; mRNA passes through a tunnel between them; A, P and E sites.

Q4. Give the functions of peroxisome. (Short)

Oxidases make H₂O₂ and catalase breaks it; detoxification of alcohol and phenols; β-oxidation of very long-chain fatty acids; plasmalogen and bile-acid synthesis; photorespiration and glyoxylate cycle in plants.

Q5. Differentiate 70S and 80S ribosomes. (Short)

70S (50S + 30S; 23S, 5S, 16S rRNA) in prokaryotes, mitochondria and chloroplasts; 80S (60S + 40S; 28S, 5.8S, 5S, 18S rRNA) in eukaryotic cytoplasm.

Exam tip

Learn the ETS order as a chain (NADH → I → Q → III → cyt c → IV → O₂) and draw the F₀–F₁ particle; most long answers are built on it.

Syllabus topics (RTMNU)

• 3.1 Lysosome: Structure, polymorphism and functions
• 3.2 Structure and function of Nucleus and Nuclear membrane
• 3.3 Chemical Composition and base composition of DNA and RNA
• 3.4 Chromatin Modelling: Supercoiling, Chromatin organization, structure of chromosome

3.1 Lysosome: structure, polymorphism and functions

Lysosomes were discovered by Christian de Duve (1955) (Nobel Prize 1974) while studying rat liver cells by centrifugation. They are called "suicide bags" of the cell because their enzymes can digest the cell itself.

Structure

• Small (0.2–0.8 µm), spherical sacs bounded by a single membrane; no DNA or ribosomes.
• Contain about 50–60 acid hydrolases working at pH ~4.5–5: proteases (cathepsins), nucleases, lipases, glycosidases, phosphatases, sulphatases. Acid phosphatase is the marker enzyme.
• The membrane has an H⁺-ATPase proton pump that keeps the inside acidic, and heavily glycosylated membrane proteins that protect it from its own enzymes.
• Formed from the trans face of the Golgi; enzymes reach them through the mannose-6-phosphate tag.

Polymorphism (forms of lysosome)

Lysosomes look different at different stages of activity – this is polymorphism.

1. Primary lysosome (storage granule): newly formed, enzymes inactive/unused, budded from Golgi.
2. Secondary lysosome (heterophagosome / digestive vacuole): formed when a primary lysosome fuses with a phagosome or endosome carrying outside material.
3. Autophagosome / autophagic vacuole: lysosome fuses with a vacuole containing the cell's own worn-out organelles (autophagy).
4. Residual body: secondary lysosome with undigested remains; thrown out by exocytosis or retained as lipofuscin (age pigment).

Functions

• Intracellular digestion (heterophagy) of food and bacteria (e.g. in macrophages, protozoa).
• Autophagy: removal of old organelles; provides nutrients in starvation.
• Autolysis: self-destruction of cells – e.g. regression of tadpole tail during metamorphosis, after death of cells.
• Extracellular digestion: enzymes released outside, e.g. bone resorption by osteoclasts; acrosome of sperm (a modified lysosome) dissolves egg coverings.
• Defence against pathogens; processing of hormones (thyroglobulin → thyroxine).
• Lysosomal storage diseases: missing enzyme causes material to pile up, e.g. Tay–Sachs disease (hexosaminidase A, ganglioside accumulates in nerve cells), Pompe's disease (glycogen), Gaucher's disease.

3.2 Structure and function of nucleus and nuclear membrane

The nucleus was discovered by Robert Brown (1831) in orchid cells. It is the control centre of the cell, holding the genetic material. Usually one per cell; RBCs of mammals and sieve tubes lack it; some cells are multinucleate (skeletal muscle fibre, Paramecium has a macro- and micronucleus).

Parts of the interphase nucleus

1. Nuclear envelope (nuclear membrane)
2. Nucleoplasm (karyolymph) – gel-like matrix with enzymes, nucleotides and the nuclear matrix (protein network).
3. Chromatin – DNA + histone proteins; euchromatin and heterochromatin.
4. Nucleolus – one or more dense, non-membranous bodies formed at the nucleolar organiser region (NOR) of certain chromosomes; site of rRNA synthesis and ribosome subunit assembly. Parts: fibrillar centre, dense fibrillar component and granular component.

Nuclear envelope – structure

• Double membrane: outer and inner membranes each ~7.5 nm, separated by the perinuclear space (~10–50 nm).
• Outer membrane is continuous with the RER and carries ribosomes.
• Inner membrane is lined by the nuclear lamina – a mesh of lamin intermediate filaments that supports the envelope and anchors chromatin.
• Nuclear pores: the two membranes fuse at many points forming pores (~3000–4000 per nucleus). Each is filled by a nuclear pore complex (NPC) – a huge structure of ~30 kinds of proteins (nucleoporins), with eight-fold (octagonal) symmetry: cytoplasmic ring, nuclear ring, central channel (~9 nm for free diffusion; up to ~25–40 nm for active transport), cytoplasmic filaments and a nuclear basket.

Functions

• Stores hereditary information (DNA) and passes it to daughter cells by replication and cell division.
• Controls all cell activities by transcription – synthesis of mRNA, tRNA, rRNA.
• Nucleolus makes ribosomes.
• Nuclear envelope separates transcription (nucleus) from translation (cytoplasm) and controls two-way traffic through pores: proteins with a nuclear localisation signal (NLS) enter with importins; RNAs and ribosomal subunits leave with exportins (Ran-GTP system).
• The envelope breaks down in prophase (lamins phosphorylated) and re-forms in telophase.

3.3 Chemical composition and base composition of DNA and RNA

Nucleic acids are polynucleotides. Discovered as "nuclein" by Friedrich Miescher (1869). Each nucleotide = nitrogenous base + pentose sugar + phosphate. Base + sugar alone = nucleoside.

Components

• Sugar: deoxyribose in DNA (no –OH at 2′ carbon); ribose in RNA.
• Purines (double ring): adenine (A), guanine (G).
• Pyrimidines (single ring): cytosine (C), thymine (T, only DNA), uracil (U, only RNA).
• Phosphodiester bond joins the 3′-OH of one sugar to the 5′-phosphate of the next, forming the sugar-phosphate backbone with 5′ → 3′ polarity. The base is joined to C1′ of the sugar by an N-glycosidic bond.

Base composition – Chargaff's rules (Erwin Chargaff, 1950)

1. In double-stranded DNA, A = T and G = C.
2. Total purines = total pyrimidines: A + G = T + C.
3. The ratio (A + T)/(G + C) varies from species to species but is constant within a species (base composition is species-specific).

Example: if a DNA has 20% adenine, then T = 20%, and G + C = 60%, so G = 30% and C = 30%.

DNA double helix – Watson and Crick (1953)

• Based on X-ray diffraction data of Rosalind Franklin and Maurice Wilkins.
• Two antiparallel strands (5′→3′ and 3′→5′) coiled round a common axis as a right-handed helix.
• Bases pair inside by hydrogen bonds: A=T (2 bonds), G≡C (3 bonds); so GC-rich DNA melts at a higher temperature.
• B-DNA: diameter 2 nm (20 Å); one turn 3.4 nm with 10 base pairs; distance between base pairs 0.34 nm; has major and minor grooves. Other forms: A-DNA (right-handed, 11 bp/turn), Z-DNA (left-handed, zig-zag, 12 bp/turn).

RNA

• mRNA (~3–5%): carries the genetic code (codons) from DNA to ribosome.
• tRNA (~10–15%): smallest (~4S, 73–93 nucleotides), clover-leaf shape (Holley), carries amino acids; has anticodon loop and CCA 3′ end.
• rRNA (~80%): most abundant; forms ribosomes with proteins.

3.4 Chromatin modelling: supercoiling, chromatin organisation, structure of chromosome

The DNA of one human cell is about 2 m long but must fit into a nucleus ~6 µm wide. This is done by step-wise packing with proteins.

Chromatin – types

• Euchromatin: lightly stained, loosely packed, genetically active (transcribed).
• Heterochromatin: darkly stained, tightly packed, mostly inactive. Constitutive (always condensed, e.g. centromere, telomere) and facultative (condensed in some cells, e.g. the Barr body – inactive X chromosome in female cells).

Levels of organisation (nucleosome model – Kornberg, 1974)

1. DNA double helix – 2 nm.
2. Nucleosome ("beads on a string") – 11 nm fibre. Core = histone octamer (2 each of H2A, H2B, H3, H4) with ~146–147 bp of DNA wound 1.65 (about 1¾) turns around it. Linker DNA (~20–60 bp) joins nucleosomes; histone H1 binds at entry/exit point. ~200 bp per nucleosome repeat. Histones are basic proteins rich in lysine and arginine (positively charged) so they bind negatively charged DNA.
3. 30 nm fibre (solenoid) – nucleosomes coiled with about 6 per turn, stabilised by H1.
4. Looped domains – 30 nm fibre forms loops (~300 nm) attached to a protein scaffold (non-histone proteins, topoisomerase II).
5. Further coiling – ~700 nm condensed section, and finally the metaphase chromatid ~1400 nm wide (≈10,000-fold compaction overall).

Supercoiling

• Extra twisting of the DNA double helix upon itself.
• Negative supercoiling (under-wound) – usual in cells; helps strand separation for replication and transcription.
• Positive supercoiling (over-wound) – builds up ahead of the replication fork.
• Controlled by topoisomerases: Type I cuts one strand; Type II (e.g. DNA gyrase in bacteria) cuts both strands and introduces negative supercoils.
• Wrapping of DNA on nucleosomes also produces supercoiling (toroidal supercoil).

Structure of a metaphase chromosome

• Chromatids: two identical sister chromatids joined at the centromere.
• Centromere (primary constriction) with kinetochore – disc-like protein where spindle fibres attach.
• Secondary constriction: may carry the NOR; distal part beyond it is the satellite (SAT-chromosome).
• Telomeres: protective ends with repeats (TTAGGG in humans); prevent ends sticking together.
• Chromomeres: bead-like thickenings along the chromonema.
• Types by centromere position: metacentric (middle, V-shape at anaphase, equal arms), submetacentric (slightly away, L-shape), acrocentric (near one end, J-shape, very short p arm), telocentric (at the end, I/rod-shape; not found in humans).
• Short arm = p, long arm = q.
• Special chromosomes: polytene (salivary gland) chromosomes of Drosophila/Chironomus larvae (Balbiani 1881) with bands and puffs; lampbrush chromosomes in amphibian oocytes.

Likely exam questions

Q1. Describe the structure and functions of the nucleus with special reference to the nuclear membrane. (Long)

• Parts: nuclear envelope, nucleoplasm, chromatin, nucleolus.
• Double membrane, perinuclear space, continuity with RER, nuclear lamina.
• Nuclear pore complex: nucleoporins, eight-fold symmetry, import/export through NLS, importins, exportins.
• Functions: heredity, transcription, ribosome synthesis, control of the cell.

Q2. Explain the organisation of chromatin into a chromosome (nucleosome model). (Long)

• 2 nm DNA → 11 nm nucleosome (histone octamer, ~146 bp, H1, linker DNA) → 30 nm solenoid → looped domains on scaffold → metaphase chromatid 1400 nm.
• Euchromatin and heterochromatin; supercoiling and topoisomerases.
• Labelled diagram of a metaphase chromosome and types by centromere position.

Q3. What is polymorphism of lysosomes? (Short)

Lysosomes take different forms at different stages: primary lysosome, secondary lysosome (heterophagosome), autophagic vacuole and residual body.

Q4. State Chargaff's rules. (Short)

In double-stranded DNA A = T and G = C; purines = pyrimidines (A + G = T + C); the ratio (A + T)/(G + C) is constant for a species but differs between species.

Q5. What is supercoiling of DNA? (Short)

Extra twisting of the double helix upon itself. Negative supercoils (under-wound) aid strand opening; positive supercoils build ahead of the replication fork; topoisomerases (e.g. DNA gyrase) control them.

Exam tip

In nucleosome questions always give the numbers (8 histones, ~146 bp, 1¾ turns, 11 nm, 30 nm, 1400 nm) and a stepwise diagram.

Syllabus topics (RTMNU)

• 4.1 Cell cycle and its regulation, Apoptosis
• 4.2 Cell Division: Mitosis and Meiosis
• 4.3 Signal transduction: Intracellular signalling, cell surface receptors, G-protein coupled receptors
• 4.4 Cellular aging and cell death

4.1 Cell cycle and its regulation, apoptosis

The cell cycle is the ordered series of events by which a cell grows, copies its DNA and divides into two daughter cells. Term given by Howard and Pelc (1953). A typical human cell in culture takes about 24 hours; yeast about 90 minutes.

Phases

G₁ + S + G₂ = interphase (~95% of cycle time); it is not a "resting stage" but a very active phase.

Regulation of the cell cycle

• Driven by cyclins (proteins whose level rises and falls) and cyclin-dependent kinases (Cdks) (enzymes always present but active only when bound to cyclin). Discovered by Hartwell, Hunt and Nurse (Nobel Prize 2001).
• G₁ cyclin–Cdk (cyclin D–Cdk4/6, cyclin E–Cdk2) pushes the cell into S; cyclin A–Cdk2 in S; cyclin B–Cdk1 = MPF (maturation/M-phase promoting factor) starts mitosis.
• Cyclins are destroyed by the anaphase-promoting complex (APC/C) via ubiquitin, allowing exit from mitosis.
• Checkpoints:
  • G₁/S checkpoint (restriction point, "START" in yeast): checks cell size, nutrients, growth factors and DNA damage. Protein Rb holds back E2F; Cdk phosphorylates Rb to release it. p53 stops the cycle (via p21) if DNA is damaged.
  • G₂/M checkpoint: checks DNA replication complete and undamaged.
  • Metaphase (spindle) checkpoint: checks all kinetochores are attached to the spindle before anaphase.
• Loss of control (mutated proto-oncogenes → oncogenes, or loss of tumour suppressors like p53, Rb) leads to cancer.

Apoptosis (programmed cell death)

• Term given by Kerr, Wyllie and Currie (1972) (Greek: "falling off", like leaves from a tree).
• A neat, genetically controlled suicide of unwanted or damaged cells without inflammation.
• Steps: cell shrinks → chromatin condenses (pyknosis) → DNA cut into ~180–200 bp fragments (ladder on gel) → nucleus breaks (karyorrhexis) → membrane blebbing → apoptotic bodies → eaten by phagocytes (recognised by phosphatidylserine on outer surface).
• Enzymes: caspases (cysteine proteases).
• Intrinsic (mitochondrial) pathway: stress/DNA damage → Bax/Bak open mitochondria → cytochrome c released → binds Apaf-1 → apoptosome → caspase-9 → caspase-3. Bcl-2 blocks it.
• Extrinsic (death receptor) pathway: Fas ligand or TNF binds death receptor → caspase-8 → caspase-3.
• Examples: separation of fingers/toes in embryo (web cells die), loss of tadpole tail, removal of self-reactive T cells in thymus, shedding of endometrium.

4.2 Cell division: mitosis and meiosis

Mitosis (equational division)

Term by Walther Flemming (1882). One diploid cell → two identical diploid daughter cells. Occurs in somatic cells for growth, repair and asexual reproduction.

1. Prophase: chromatin condenses into visible chromosomes, each with two sister chromatids; centrosomes move to opposite poles and form the spindle; nucleolus and nuclear envelope disappear (late prophase/prometaphase).
2. Metaphase: chromosomes most condensed; spindle fibres attach to kinetochores; chromosomes line up at the equatorial (metaphase) plate. Best stage to study chromosome morphology and count.
3. Anaphase: centromeres split; sister chromatids separate and move to opposite poles (centromere first, arms trailing → V, L, J shapes).
4. Telophase: chromosomes reach poles and decondense; nuclear envelope and nucleolus re-form – two nuclei.
5. Cytokinesis: in animal cells a cleavage furrow forms by an actin–myosin contractile ring (centripetal); in plant cells a cell plate forms from Golgi vesicles (centrifugal).

Significance: keeps chromosome number constant, growth, repair and replacement of cells (skin, gut lining, blood cells), regeneration, asexual reproduction.

Meiosis (reduction division)

Term by Farmer and Moore (1905). One diploid (2n) cell → four haploid (n) cells, genetically different. Occurs in germ cells during gamete formation (gametogenesis in animals). Two divisions with only one DNA replication.

Meiosis I (heterotypic – reduces chromosome number)

• Prophase I – long and complex, five sub-stages:
  1. Leptotene: chromosomes become visible as long thin threads.
  2. Zygotene: homologous chromosomes pair (synapsis) with the help of the synaptonemal complex; a pair = bivalent.
  3. Pachytene: each bivalent shows four chromatids (tetrad); crossing over between non-sister chromatids, enzyme recombinase; recombination nodules.
  4. Diplotene: synaptonemal complex dissolves; homologues start separating but stay joined at chiasmata (X-shaped points). Can last years in human oocytes (dictyotene).
  5. Diakinesis: chiasmata move to ends (terminalisation); nucleolus and nuclear envelope disappear; spindle forms.
• Metaphase I: bivalents line up at the equator (two rows of chromosomes).
• Anaphase I: homologous chromosomes separate (each still with two chromatids) – this is the actual reduction; centromeres do not split.
• Telophase I + cytokinesis → two haploid cells (dyad). A short interkinesis follows with no DNA replication.

Meiosis II (homotypic – like mitosis)

• Prophase II, metaphase II, anaphase II (centromeres split, sister chromatids separate), telophase II → four haploid cells (tetrad).

Significance of meiosis

• Keeps chromosome number constant from generation to generation (gametes n + n → zygote 2n).
• Crossing over and independent assortment of chromosomes create genetic variation – raw material for evolution.
• Errors (non-disjunction) cause aneuploidy, e.g. Down syndrome (trisomy 21).

4.3 Signal transduction: intracellular signalling, cell surface receptors, G-protein coupled receptors

Signal transduction is the process by which a cell receives an outside signal (first messenger) and converts it into a response inside the cell. Steps: reception → transduction (relay and amplification) → response.

Types of signalling by distance

• Endocrine: hormones travel in blood to distant targets (insulin).
• Paracrine: acts on nearby cells (growth factors, histamine).
• Autocrine: cell responds to its own signal (interleukin-2 on T cells).
• Synaptic: neurotransmitter across a synapse.
• Juxtacrine/contact-dependent: through membrane molecules or gap junctions.

Intracellular signalling

• Intracellular (nuclear) receptors: small lipid-soluble signals – steroid hormones (oestrogen, testosterone, cortisol), thyroxine, vitamin D, retinoic acid – cross the membrane and bind receptors in cytoplasm or nucleus; the hormone–receptor complex binds DNA and switches genes on/off.
• Second messengers carry the signal inside after a surface receptor is activated: cAMP (Earl Sutherland, Nobel 1971), cGMP, IP₃, DAG, Ca²⁺, nitric oxide (NO).
• Protein kinases (PKA, PKC, MAP kinases) add phosphate to target proteins in a cascade that amplifies the signal; phosphatases switch it off.

Cell surface receptors – three main classes

1. Ion-channel-linked receptors (ligand-gated channels): binding opens a channel, e.g. nicotinic acetylcholine receptor at the neuromuscular junction (Na⁺ enters).
2. G-protein coupled receptors (GPCRs) – see below.
3. Enzyme-linked receptors, mainly receptor tyrosine kinases (RTKs): ligand (insulin, EGF, NGF) binding causes two receptors to pair (dimerise) and phosphorylate each other's tyrosines; this activates Ras → Raf → MEK → MAP kinase pathway leading to cell growth and division.

G-protein coupled receptors (GPCRs)

• Largest family of surface receptors; single polypeptide that crosses the membrane seven times (7-transmembrane / serpentine receptors). Examples: β-adrenergic receptor (adrenaline), rhodopsin (light), olfactory receptors, glucagon receptor.
• G-protein = heterotrimer of α, β and γ subunits; the α subunit binds GDP (inactive) or GTP (active).
• Mechanism:
  1. Ligand binds GPCR → receptor changes shape.
  2. Receptor makes Gα release GDP and bind GTP.
  3. Gα-GTP separates from βγ and activates an effector enzyme.
  4. Gs → adenylyl cyclase → ATP to cAMP → protein kinase A (PKA) → response (e.g. glycogen breakdown in liver by adrenaline/glucagon). Gi inhibits adenylyl cyclase.
  5. Gq → phospholipase C → splits PIP₂ into IP₃ (releases Ca²⁺ from ER) and DAG (activates protein kinase C).
  6. Gα slowly hydrolyses GTP to GDP (built-in timer) and rejoins βγ → switched off; phosphodiesterase breaks cAMP to AMP.
• Diseases: cholera toxin locks Gs in the active state (huge cAMP → water loss, diarrhoea); pertussis toxin blocks Gi.

4.4 Cellular ageing and cell death

Cellular ageing (senescence)

• Gradual decline in the structure and function of cells with time, ending in loss of the ability to divide.
• Hayflick limit (Leonard Hayflick, 1961): normal human fibroblasts in culture divide only about 40–60 times and then stop (replicative senescence).
• Signs: larger flattened cells, fewer and abnormal mitochondria, accumulation of lipofuscin (age pigment) in residual bodies, cross-linked proteins (collagen), damaged DNA, slower repair, weaker membranes.

Theories of ageing

1. Telomere shortening theory: telomeres shorten by 50–200 bp at each division because DNA polymerase cannot copy the very end (end-replication problem); short telomeres trigger arrest. Telomerase (reverse transcriptase with RNA template; Blackburn, Greider and Szostak – Nobel 2009) is active in germ cells, stem cells and cancer cells, keeping them dividing.
2. Free radical theory (Denham Harman, 1956): reactive oxygen species (superoxide O₂⁻, hydroxyl radical ·OH, H₂O₂) from mitochondria damage DNA, proteins and lipids. Defence: superoxide dismutase, catalase, glutathione peroxidase, vitamins C and E.
3. Mitochondrial theory: mutations pile up in mitochondrial DNA → less ATP, more free radicals.
4. Somatic mutation / DNA damage theory: errors accumulate faster than repair.
5. Cross-linkage theory: proteins like collagen get cross-linked (e.g. by glucose – glycation) → stiff tissues, wrinkles.
6. Programmed (genetic clock) theory: lifespan set by genes; e.g. progeria and Werner syndrome (premature ageing gene defects).

Cell death – types

• Autophagic cell death: excessive self-digestion by lysosomes (autophagosomes).
• Balance of cell division and cell death keeps tissue size constant (homeostasis); too little death → cancer, too much → degenerative disease (Alzheimer's, Parkinson's).

Likely exam questions

Q1. Describe the cell cycle and its regulation. (Long)

• Phases G₁, S, G₂, M and G₀; interphase is active.
• Cyclins and Cdks; MPF (cyclin B–Cdk1); APC/C.
• Checkpoints G₁/S, G₂/M, metaphase; role of Rb and p53; failure leads to cancer.

Q2. Describe meiosis with special reference to prophase I. Add its significance. (Long)

• Leptotene, zygotene (synapsis, synaptonemal complex), pachytene (crossing over), diplotene (chiasmata), diakinesis (terminalisation).
• Metaphase I, anaphase I (homologues separate), telophase I, meiosis II.
• Significance: constant chromosome number, variation; comparison with mitosis.

Q3. What is apoptosis? (Short)

Programmed cell death: cell shrinks, chromatin condenses, DNA is cut into ladder fragments, membrane blebs into apoptotic bodies eaten by phagocytes; caspases carry it out; no inflammation. Example: loss of tadpole tail.

Q4. Explain the mechanism of G-protein coupled receptor. (Short)

A 7-transmembrane receptor; ligand binding makes Gα swap GDP for GTP, Gα separates from βγ and activates adenylyl cyclase (cAMP → PKA) or phospholipase C (IP₃ + DAG); GTP hydrolysis switches it off.

Q5. What is the Hayflick limit? (Short)

Normal human cells in culture divide only about 40–60 times before they stop (replicative senescence), mainly because telomeres shorten at each division.

Exam tip

For meiosis, write the five prophase I sub-stages in order with one key event each; the mnemonic "Lazy Zebras Prefer Dry Days" helps.