Cytoskeleton, Cellular Motility, and Extracellular Matrix: Study Notes (Chapters 13–15)

Cytoskeletal Filaments: Structure and Comparison

The Three Major Cytoskeletal Filaments

The cytoskeleton is composed of three primary filament systems, each with distinct structures, subunits, and cellular functions. Understanding their differences is essential for grasping cell architecture and dynamics.

Feature	Microtubules (MTs)	Microfilaments (Actin / MFs)	Intermediate Filaments (IFs)
Diameter	25 nm (thickest)	~7 nm (thinnest)	8–12 nm (intermediate)
Subunit	alpha/\beta-tubulin heterodimer	G-actin monomer	Diverse fibrous proteins (keratins, vimentin, lamins)
Nucleotide	GTP (\beta-tubulin)	ATP	None
Polarity	Polar (+ and - ends)	Polar (barbed + / pointed -)	Nonpolar (symmetrical tetramers)
Motors	Kinesins (+), Dyneins (-)	Myosins (mostly +)	None
Primary Origin	MTOC / Centrosome (\gamma-TuRC)	Cell cortex, Arp2/3, formins	Desmosomes, hemidesmosomes, nuclear lamina
Core Functions	Organelle transport, mitotic spindle, cilia/flagella	Muscle contraction, cytokinesis, cell cortex shape	Mechanical tensile strength, nuclear lamina stability

Chapter 13: Cytoskeletal Mechanics & Kinetics

Microtubule Structure and Dynamics

• Structural Polarity: Microtubules are composed of 13 protofilaments aligned in parallel, imparting polarity. The (+) end (exposed \beta-tubulin) is fast-growing and dynamic, while the (–) end (exposed \alpha-tubulin) is slow-growing and typically anchored at the microtubule organizing center (MTOC).

• MTOC (Centrosome): Located near the nucleus, the MTOC anchors and caps only the minus ends of microtubules via \gamma-tubulin ring complexes. Inhibition of \gamma-tubulin halts de novo nucleation.

• Kinetics & Assembly:

  • Lag Phase: The nucleation step is rate-limiting; adding pre-formed seeds eliminates this lag.

  • Critical Concentration (Cc): The free subunit concentration at which assembly and disassembly rates are equal.

  • Treadmilling: At Cc, subunits add at the (+) end and leave at the (–) end, resulting in no net change in polymer length.

• Mitotic Drugs & Regulators:

  • Catastrophins (e.g., MCAK/Kinesin-13): Destabilize microtubules at kinetochores and spindle poles, promoting chromosome segregation.

  • Taxol (Paclitaxel): Stabilizes microtubules, preventing disassembly and blocking mitosis; used in cancer therapy.

Actin Dynamics & Regulatory Proteins

• Evolutionary Conservation: Actin is highly conserved (~90% identity across eukaryotes).

• Non-Cytoskeletal Function: Actin does not participate in passive transport, which does not require cytoskeletal tracks or ATP.

• Key Regulators:

  • Profilin: Promotes ATP exchange on G-actin, enhancing (+) end growth.

  • Cofilin: Binds ADP-actin near the (–) end, severs filaments, and accelerates turnover.

  • Gelsolin: Severs actin networks and caps (+) ends, liquefying the cortical gel.

  • Fimbrin & Villin: Cross-link parallel actin bundles, supporting microvilli structure.

• Rho GTPase Signals:

  • Rho: Induces stress fibers and cytokinetic cleavage furrow formation.

  • Rac: Promotes lamellipodia (branched actin meshwork).

  • Cdc42: Drives filopodia (actin bundles/spikes).

• Exam Trap: Nuclear envelope breakdown is driven by lamin (IF) phosphorylation, not by Rho or microfilaments.

Intermediate Filaments & Associated Diseases

• Assembly Sequence: Monomer $\rightarrow$ Coiled-coil Dimer $\rightarrow$ Antiparallel Tetramer $\rightarrow$ Protofilament $\rightarrow$ 8–12 nm IF.

• Lack of Polarity: Due to antiparallel tetramer assembly, IFs are nonpolar and cannot direct motor transport.

• Epidermolysis Bullosa Simplex (EBS): Point mutations in keratins (K5/K14) weaken tensile strength, causing skin blistering under mild stress.

• FtsZ & Endosymbiosis: FtsZ is a bacterial tubulin homolog forming the contractile ring in binary fission and organelle division, supporting the endosymbiotic origin of mitochondria and chloroplasts.

Chapter 14: Cellular Motility & Motor Systems

Intracellular Transport and Motor Proteins

• Dynein + Dynactin: Minus-end-directed motor; mediates retrograde transport (ER to Golgi, periphery to MTOC).

• Kinesin-1: Plus-end-directed motor; mediates anterograde transport (MTOC to periphery, e.g., ER extension).

• Kinesin Stepping Mechanism:

  • One head remains bound (high affinity) while the other steps (low affinity), ensuring processivity.

• Energy Conversion: Motor proteins convert ATP hydrolysis energy into mechanical work.

Cilia and Axonemal Motility

• Axoneme Pattern: "9 + 2" arrangement (nine outer doublets around two central singlets); basal bodies have a "9 + 0" triplet structure.

• A vs. B Tubules:

  • A Tubule: 13 protofilaments (complete); anchors dynein arms, nexin links, and radial spokes.

  • B Tubule: 10–11 protofilaments (incomplete); serves as the dynamic track for dynein movement.

• Sliding vs. Bending:

  • With intact radial spokes/nexin links, dynein walking causes ciliary bending.

  • Without these links, doublets slide past each other without bending.

• Nucleotide Rule: Axoneme structure requires GTP; dynein motility requires ATP.

Skeletal Muscle & The Cross-Bridge Cycle

• Sarcomere Bands:

  • I Band: Light; thin filaments only; shortens during contraction.

  • A Band: Dark; spans thick filaments; length remains constant.

  • Titin: Anchors thick filaments to the Z disc, preventing overstretching.

• Cross-Bridge Cycle Steps:

  1. Formation: Myosin head binds actin.

  2. Power Stroke: Pi release triggers neck pivot, pulling thin filament toward M line; ADP releases.

  3. Dissociation: ATP binding causes myosin detachment from actin.

  4. Cocking: ATP hydrolysis resets myosin to the high-energy state.

• Excitation & Regulation:

  • Acetylcholine (ACh) binds sarcolemma receptors, triggering depolarization.

  • Ca2+ is released from the sarcoplasmic reticulum, binds Troponin C, and moves tropomyosin to allow contraction.

  • Sarin inhibits acetylcholinesterase, causing continuous depolarization and muscle spasms.

  • Cardiac muscle cells are single, branched, and joined by intercalated discs/gap junctions; not multinucleated syncytia.

Smooth Muscle & Non-Muscle Motility

• Smooth Muscle Regulation: Lacks troponin; Ca2+ binds calmodulin, activating MLCK, which phosphorylates myosin heads to trigger contraction. MLCK inhibitors halt contraction.

• Amoeboid Gel–Sol Transition: Gelsolin, activated by Ca2+, severs actin at the pseudopodium base, allowing endoplasm to flow forward.

• Chemotaxis: Receptor-mediated, ATP-dependent movement toward chemical signals; does not require new gene expression.

Chapter 15: Cell Junctions & The Extracellular Matrix (ECM)

Cell Junctions: Types and Functions

Junction Type	Transmembrane Proteins	Cytoskeletal Track	Core Function
Tight Junction	Claudins, Occludins	Actin microfilaments	Gate (blocks paracellular leaks) and Fence (maintains apical/basolateral polarity)
Desmosome	Desmoglein, Desmocollin	Intermediate Filaments	Resists mechanical shearing; abundant in skin
Hemidesmosome	\alpha_6\beta_4 Integrin	Intermediate Filaments	Anchors basal epithelial cells to basal lamina
Gap Junction	Connexins (connexon)	None	Direct intercellular communication; no adhesive role

• Intracellular Plaque: Dense protein complex (e.g., desmoplakin, catenins) links adhesion receptors to the cytoskeleton.

Extracellular Matrix (ECM) Components & Clinical Conditions

• Fibroblasts: Main connective tissue cells producing procollagen and ECM precursors.

• Collagen Defects:

  • Scurvy: Vitamin C deficiency impairs prolyl/lysyl hydroxylases, leading to unstable collagen, fragile capillaries, and poor healing.

  • Ehlers-Danlos Syndrome: Genetic defects in collagen or processing enzymes cause hypermobile joints and fragile skin.

  • Type V Collagen: Co-assembles with Type I collagen in the dermis.

• Basal Lamina & Cancer: Composed of Type IV collagen and laminins; carcinoma cells secrete MMPs (Type IV collagenases) to breach the basal lamina during metastasis.

• Non-Collagenous ECM:

  • Fibronectin: Multiadhesive glycoprotein with RGD sequence; binds integrins and fibrin, stabilizing blood clots.

  • Hyaluronate (Hyaluronic Acid): Large, non-sulfated GAG that retains water for joint lubrication.

  • Pectins: Galacturonic-acid-rich polysaccharides in plant cell walls; absent in animals.

  • ECM Non-Role: Integrin signaling regulates migration, polarity, and survival, but not nuclear DNA replication.

High-Frequency Exam Traps & Flash Facts

• MTOC Anchoring: MTOC stabilizes and caps the (–) end, leaving the (+) end dynamic.

• Dynein vs. Kinesin: Dynein moves ER to Golgi (minus-directed); Kinesin moves Golgi to ER/periphery (plus-directed).

• IF Symmetry: IFs are nonpolar due to antiparallel tetramer assembly; lack motor proteins.

• EBS vs. Scurvy: EBS is an IF (keratin) defect; Scurvy is an ECM (collagen) defect.

• Sarcomere Changes: During contraction, I band and H zone narrow; A band remains constant.

• Axoneme Digestion: Removing radial spokes prevents bending, causing sliding of doublets.

• Gap Junction Role: Connexins form gap junctions for solute communication, not adhesion.

• Basal Lamina Target: Tumors degrade Type IV collagen and laminin (not Type I) to invade tissues.