Chapter 13: Cytoskeletal Systems

Cytoskeleton Overview

The cytoskeleton is a dynamic, three-dimensional network of protein filaments present in both prokaryotic and eukaryotic cells. It provides structural support, internal organization, and enables cell movement and division.

• Determines and maintains cell shape

• Organizes cytoplasm and positions organelles

• Supports cell motility and division

• Moves organelles and macromolecules

Major Cytoskeletal Elements in Eukaryotes

• Microtubules: Hollow tubes (~25 nm diameter), composed of α- and β-tubulin heterodimers.

• Microfilaments: Thin filaments (~7 nm diameter), polymers of actin (G-actin monomers assemble into F-actin).

• Intermediate Filaments: Diameter 8–12 nm, built from various IF proteins (e.g., keratins, vimentin, neurofilaments).

Bacterial Cytoskeletal Elements

• MreB (actin-like): Maintains rod shape, associates with cell wall synthesis.

• FtsZ (tubulin-like): Forms Z ring at division site, marks cytokinesis.

• Crescentin (intermediate filament-like): Maintains curved cell shape.

Microtubule Structure and Types

• Subunit: Tubulin heterodimer (α-tubulin + β-tubulin).

• Protofilaments: Linear chains of tubulin dimers; 13 protofilaments form a microtubule.

• Polarity: Plus (+) and minus (−) ends; crucial for directed growth and motor protein movement.

• Singlet: 13-protofilament tube.

• Doublet: 13-protofilament A tubule + incomplete B tubule.

• Triplet: A tubule + two incomplete B and C tubules.

Microtubule Assembly

Microtubules assemble by reversible polymerization of tubulin dimers, a process called microtubule nucleation and elongation.

• Lag phase (nucleation): Formation of oligomers and protofilaments.

• Elongation phase: Rapid growth by addition of tubulin dimers.

• Plateau phase: Balance between assembly and disassembly.

Factors affecting kinetics: tubulin concentration, GTP/Mg2+ availability, temperature.

Microtubule Treadmilling

Treadmilling is a dynamic process where tubulin dimers add at the plus end and are lost at the minus end, maintaining constant length but moving subunits through the filament.

Microtubule Drugs

• Colchicine: Binds β-tubulin, inhibits polymerization, destabilizes microtubules.

• Nocodazole: Binds β-tubulin, inhibits assembly, reversible effects.

• Paclitaxel (Taxol): Stabilizes microtubules, prevents disassembly, arrests mitosis.

Dynamic Instability and GTP Cap

Microtubules exhibit dynamic instability: some grow, others shrink. The GTP cap at the plus end stabilizes growth; loss of the cap leads to rapid depolymerization (catastrophe).

Microtubule-Organizing Centers (MTOCs)

• Centrosome: Main MTOC in animal cells.

• Centrioles: Cylindrical structures within centrosome.

• Pericentriolar material: Contains γ-tubulin ring complexes (γ-TuRCs) for nucleation.

γ-Tubulin

• Found in MTOCs, not along microtubules.

• Essential for nucleation and anchoring minus ends.

Microtubule Polarity from MTOC

• Minus ends anchored at MTOC; plus ends grow outward.

Microtubule-Associated Proteins (MAPs)

• MAPs bind microtubules, crosslink them, and connect to other structures.

• Abnormal MAPs (e.g., Tau) can cause neurofibrillary tangles (Alzheimer's).

Microtubule Stabilizing and Destabilizing Proteins

• Stabilizing: MAPs, +TIP proteins (EB1).

• Destabilizing/Severing: Stathmin/Op18, catastrophins (MCAK), katanin.

Microfilaments: Actin

• G-actin: Globular monomer, binds ATP/ADP.

• F-actin: Polymerized, double-stranded helix, polar (barbed plus end, pointed minus end).

Motility Structures in Crawling Cells

• Lamellipodia: Sheet-like, branched actin network.

• Filopodia: Finger-like, parallel actin bundles.

Actin Assembly Regulation

• Controlled by actin-binding proteins, phospholipids, and Rho family GTPases (Rho, Rac, Cdc42).

• Rho: stress fibers; Rac: lamellipodia; Cdc42: filopodia.

Intermediate Filaments: Classes and Structure

• Six classes: Keratins (I-II), Vimentin-family (III), Neurofilaments (IV), Nuclear lamins (V), Nestin (VI).

• Diameter: 8–12 nm (smaller than microtubules, larger than microfilaments).

Chapter 15: Cell Adhesions, Junctions, and Extracellular Structures

Types of Animal Tissues

• Epithelial tissue: Compact, polarized sheets, attached to basal lamina.

• Connective tissue: Loosely organized, embedded in ECM.

Cell-Cell Junctions

• Adhesive junctions: Adherens (actin, cadherins), desmosomes (intermediate filaments, cadherins).

• Tight junctions: Seal epithelial layers, control permeability.

• Gap junctions: Connexons form channels for communication.

• Plant cells: Plasmodesmata (analogous to gap junctions).

Homophilic vs. Heterophilic Interactions

• Homophilic: Same molecule binds across cells (e.g., cadherins).

• Heterophilic: Different molecules bind (e.g., selectins, IgSF proteins).

Cell-Cell Adhesive Junctions: Proteins

• Adherens junctions: Cadherins, actin, linker proteins.

• Desmosomes: Cadherins, intermediate filaments, attachment proteins.

Cancer Cell Metastasis

• Loss of cell-cell adhesion (E-cadherin), increased motility (Rho GTPases), secretion of proteases, blood flow/organ factors.

Other Cell Adhesion Molecules

• IgSF proteins: Flexible adhesion, immune/neural functions.

• Selectins: Carbohydrate-binding, leukocyte rolling.

• Membrane glycoproteins: Cell recognition markers.

Tight Junctions in Epithelial Cells

• Seal layers, control permeability, partition membrane domains.

Gap Junctions vs. Tight Junctions

• Tight junctions: Seal, no open channels.

• Gap junctions: Open channels (connexons), direct communication.

Extracellular Matrix (ECM): Types and Molecules

Three types of ECM:

• Bone (rigid), cartilage (flexible), loose connective tissue (soft, fibrous).

Three classes of ECM molecules:

Collagen: Most Abundant ECM Component

• Provides mechanical strength, forms fibers, most abundant protein in vertebrates.

Collagen Assembly

• Triple helix (procollagen) formed in ER.

• Procollagen peptidase removes extensions after secretion.

• Collagen molecules assemble into fibrils and fibers.

Elastin: ECM Stretching

• Forms elastic fibers, crosslinked network, allows tissues to stretch and recoil.

Proteoglycans

• Core protein + GAG chains, highly hydrated, gel-like matrix, tissue resilience.

Fibronectin: Cell Movement and Cancer

• Adhesive glycoprotein, bridges cells and ECM, guides migration, altered in cancer progression.

Basal Lamina

• Structural support, permeability barrier, selective filter under epithelia.

Integrins and Hemidesmosomes

• Integrins: α/β heterodimers, bind ECM outside, cytoskeleton inside.

• Hemidesmosomes: α6β4 integrin binds laminin, connects to intermediate filaments (keratin).

Anchorage-Dependent Growth and Apoptosis

• Cells require ECM attachment via integrins to grow; loss triggers apoptosis.

• Cancer cells can grow anchorage-independently.

Dystrophin: Muscle Cell Attachment

• Links muscle cells to ECM, stabilizes during contraction; defects cause muscular dystrophy.

Plant vs. Animal Structural Framework

• Plant cells: rigid cell wall (cellulose, hemicellulose, pectin, glycoproteins).

• Animal cells: flexible ECM.

Primary vs. Secondary Cell Wall

Plasmodesmata: Plant Gap Junction Analog

• Cytosolic channels through cell wall, direct communication between plant cells.

Chapter 24: The Cell Cycle and Mitosis

Cell Division: Mitosis vs. Cytokinesis

• Mitosis: Nuclear division, chromosome segregation.

• Cytokinesis: Cytoplasmic division, formation of two cells.

Cell Cycle Stages and Timing

• G1: Growth, metabolism, decision to divide.

• S: DNA replication.

• G2: Preparation for division.

• M: Mitosis and cytokinesis.

Interphase (G1, S, G2) ~95% of cycle; M phase ~30–45 min.

Cell Cycle Measurement Techniques

• Mitotic index (microscopy)

• Flow cytometry (DNA content)

• BrdU incorporation (S-phase labeling)

Five Stages of Mitosis

• Prophase: Chromatin condenses, nucleolus disperses.

• Prometaphase: Nuclear envelope breaks, spindle attaches.

• Metaphase: Chromosomes align at metaphase plate.

• Anaphase: Sister chromatids separate, move to poles.

• Telophase: Chromosomes decondense, nuclear envelope reforms.

Mitotic Spindle Microtubule Polarity

• Minus ends at centrosomes (poles), plus ends toward cell interior/chromosomes.

Kinetochore

• Multiprotein complex at centromere, attaches spindle microtubules, ensures chromosome segregation.

Chromosome Alignment at Metaphase Plate

• Poleward pulling (dynein), anti-poleward pushing (CENP-E kinesin), congression (back-and-forth movement).

Motor Proteins in Anaphase

• Chromosome-to-pole movement (kinesins, dynein), spindle elongation (Eg5 kinesin), pole pulling (dynein).

Cytokinesis: Cleavage Furrow

• Contractile ring of actin and myosin forms cleavage furrow, pinches cell into two.

Cleavage Mechanism: Proteins Involved

• Actin filaments, nonmuscle myosin II, RhoA GTPase, centralspindlin complex.

Plant vs. Animal Cytokinesis

• Animal: Contractile ring, cleavage furrow.

• Plant: Cell plate formation, new cell wall.

Cell Cycle Variation

• Length and phase duration vary by cell type and organism; some cells exit cycle permanently (G0).

Cell Cycle Transition Points and Checkpoints

Discovery of Cytoplasmic Cell Cycle Regulators

• Cell fusion experiments showed phase-specific cytoplasmic factors drive cell cycle transitions (e.g., S-phase, mitosis).

Major Protein Regulators: Cyclins and Cdks

• Cdk–cyclin complexes phosphorylate targets to drive cell cycle transitions.

• APC/C (anaphase-promoting complex) triggers chromatid separation and mitotic exit.

Cyclin Regulation

• Cyclin levels oscillate; Cdks active only when bound to cyclins.

• APC/C degrades mitotic cyclin, inactivates Cdk, allows mitotic exit.

Mitotic Cyclins: Targets and Mechanism

• Phosphorylate proteins for nuclear envelope breakdown, chromosome condensation, spindle assembly.

• APC/C ubiquitinates securin, releases separase, cleaves cohesins, triggers anaphase.

Rb/E2F Control of G1/S Transition

• Rb binds E2F, blocks S-phase gene transcription.

• G1 Cdk–cyclin phosphorylates Rb, releases E2F, activates S-phase genes.

Licensing and Prevention of Relicensing

• Origins licensed in G1 (ORC, MCMs); Cdk and geminin prevent relicensing in S phase.

p53: Guardian of the Genome

• Activated by DNA damage, induces p21 (cell cycle arrest) or Puma (apoptosis).

Unicellular vs. Multicellular Division Control

• Unicellular: Nutrient availability.

• Multicellular: Growth factors, mitogens, extracellular signals.

Ras and PI 3-Kinase–Akt Pathways

• Ras pathway: Growth factor → Ras → Raf → MEK → MAPK → transcription factors → cell cycle genes.

• PI 3-Kinase–Akt: Growth factor → PI 3-kinase → PIP3 → Akt → Bad/Rheb/TOR → cell growth, survival.

Cell Cycle Inhibitors

• Inhibitory growth factors (TGFβ), Cdk inhibitors (p15, p21), checkpoint signals.

Apoptosis vs. Necrosis

• Apoptosis: Programmed, orderly cell death; DNA fragmentation, cell shrinkage, phagocytosis.

• Necrosis: Uncontrolled, cell swelling and rupture, damages neighbors.

Apoptosis Steps and Triggers

• Death signals (Fas/CD95), withdrawal of survival factors, DNA damage (p53).

• Initiator procaspases activated, cleave executioner procaspases, caspases dismantle cell.

Direct vs. Indirect Apoptosis

• Direct: Death receptor activation (procaspase-8).

• Indirect: Mitochondrial pathway (cytochrome c, Apaf-1, procaspase-9).

Example: In Alzheimer's disease, abnormal Tau MAPs form tangles, disrupting microtubule organization and neuronal function.

Additional info: The study guide covers foundational concepts for cytoskeletal systems, cell junctions, extracellular matrix, and cell cycle regulation, with emphasis on molecular mechanisms and clinical relevance (e.g., cancer, muscular dystrophy, apoptosis).