05 — Intermediate Filaments and Cell Migration

Prerequisite Concepts

• Actin polymerization, Arp2/3, and formins (Topic 01)
• Myosin II motor function (Topic 02)
• Microtubule organization and motor proteins (Topics 03, 04)
• Basic concepts of cell adhesion and extracellular matrix

Core Explanation

Intermediate Filaments: Structure and Properties

Intermediate filaments (IFs) are named for their diameter: ~10 nm — between actin microfilaments (~7 nm) and microtubules (~25 nm). Unlike actin and tubulin, which are globular proteins that polymerize into polarized filaments and hydrolyze nucleotides, IF proteins are fibrous with a conserved central α-helical coiled-coil rod domain flanked by variable head and tail domains.

Key structural principles:

1. Two IF polypeptides dimerize via coiled-coil interactions → parallel dimer.
2. Two dimers associate in an antiparallel, staggered fashion → tetramer (the soluble subunit).
3. Tetramers assemble end-to-end and laterally into non-polarized, rope-like filaments that do not require nucleotide binding for polymerization.

Critical distinction from actin/MTs: Intermediate filaments are apolar (antiparallel tetramer assembly) and do not bind or hydrolyze nucleotides. Their assembly is driven purely by protein–protein interactions and is regulated by phosphorylation, not NTP hydrolysis.

Major Classes and Tissue Specificity

Dynamics and Regulation

IFs are more stable than actin or microtubules (half-lives of hours vs minutes), but they are not static. Subunits exchange along the filament length (not just ends, due to apolar assembly), and phosphorylation regulates disassembly — e.g., vimentin phosphorylation by Rho-kinase during mitosis causes IF network collapse. This dynamic stability suits IFs' role as long-term mechanical scaffolds that can remodel when needed.

Nuclear Lamins

Lamins (A/C, B1, B2) form a meshwork — the nuclear lamina — on the inner face of the nuclear envelope. The lamina:

• Provides mechanical support to the nucleus.
• Anchors chromatin (via lamin-associated domains, LADs) and nuclear pore complexes.
• Disassembles during mitosis via lamin phosphorylation by CDK1.

Laminopathies — diseases caused by lamin mutations — illustrate the diverse roles of IFs:

• Hutchinson-Gilford progeria: Lamin A mutation (progerin) → nuclear fragility, premature aging phenotype.
• Emery-Dreifuss muscular dystrophy: Lamin A/C mutations → skeletal and cardiac muscle degeneration.

The Cell Migration Cycle

Cell migration is a coordinated, cyclic process of four overlapping phases:

Phase 1: Front-Rear Polarity

A migrating cell establishes asymmetry: a leading edge (front) and a trailing edge (rear). This is driven by:

• Rho-family GTPases: Rac and Cdc42 activity at the front; RhoA activity at the rear.
• PI3K signaling: PIP₃ accumulation at the leading edge recruits Rac GEFs (guanine nucleotide exchange factors), reinforcing front identity.
• Microtubule stabilization: Selectively stabilized microtubules orient toward the leading edge, delivering cargo and signaling molecules.

Phase 2: Protrusion

The leading edge extends forward via actin polymerization:

• Lamellipodia: Broad, sheet-like protrusions driven by Arp2/3-branched actin networks. Rac GTPase activates WAVE complex → Arp2/3 activation → dense, branched actin mesh that pushes the membrane forward. Lamellipodia are the primary protrusive structures of migrating fibroblasts and keratinocytes.
• Filopodia: Thin, finger-like projections containing formin-nucleated, bundled actin. Cdc42 activates formins (e.g., mDia2) and Ena/VASP proteins, producing parallel actin bundles that act as sensory probes exploring the environment.
• Invadopodia/Podosomes: Proteolytic protrusions that degrade ECM in invasive cancer cells and osteoclasts.

Membrane delivery from the endocytic/recycling pathway (via VAMP3, Rab11) supplies the lipid needed for expanding protrusions.

Phase 3: Adhesion (Focal Adhesions)

Protrusions must attach to the substrate (ECM) to generate traction. Focal adhesions are the key structures:

• Integrins: Heterodimeric (α/β) transmembrane receptors that bind ECM ligands (fibronectin, collagen, laminin). Activated integrins cluster and recruit intracellular adaptors.
• Talin: Binds integrin cytoplasmic tails and actin; activates integrins (inside-out signaling) and transmits force from the actomyosin system to the ECM.
• Vinculin: Binds talin and actin; reinforces talin–integrin linkages under force. Vinculin's autoinhibition is released by talin binding and mechanical tension.
• FAK (Focal Adhesion Kinase): A non-receptor tyrosine kinase recruited to nascent adhesions. Autophosphorylation (Y397) creates a docking site for Src kinase and downstream signaling (survival, proliferation, adhesion turnover).

Focal adhesions are force-sensitive: they grow and mature under tension from the actomyosin system. This mechanosensitivity couples adhesion strength to the contractile machinery.

Phase 4: Contraction and Rear Detachment

Actomyosin contraction — actin filaments (often in stress fibers) and nonmuscle myosin II — generates traction forces that pull the cell body forward. RhoA activates ROCK, which phosphorylates myosin regulatory light chain (activating myosin II) and inhibits myosin phosphatase.

At the rear, focal adhesions must disassemble (detachment). Mechanisms include:

• Calpain proteases: Cleave talin and FAK at the rear.
• Microtubule targeting: Microtubules grow into rear adhesions and trigger disassembly (potentially via KIF2C/MCAK-mediated FAK dephosphorylation).
• Endocytosis of integrins from the rear membrane.

If rear detachment fails while the front continues to protrude, the cell becomes stretched and cannot translocate — adhesion turnover must be coordinated with protrusion.

Rho-Family GTPases: The Conceptual Framework

Three Rho-family GTPases act as molecular switches coordinating migration:

Mutual antagonism: Rac and RhoA are often mutually inhibitory — Rac activation at the front suppresses RhoA, and RhoA at the rear suppresses Rac. This creates a self-reinforcing polarity system: Rac specifies "front" behavior (protrusion), RhoA specifies "rear" behavior (contraction).

Disease Connections

• Epidermolysis bullosa simplex: Keratin 5/14 mutations → skin fragility; basal keratinocytes rupture under mechanical stress.
• Progeria: Lamin A mutation (progerin) → nuclear fragility, premature aging.
• Metastasis: Cancer cells upregulate vimentin and switch to a mesenchymal migration program (EMT). Arp2/3, cofilin, and formin dysregulation promote invasive protrusions.
• Leukocyte adhesion deficiency: Defective integrin function → impaired immune cell migration → recurrent infections.
• Wound healing: Failed keratinocyte migration delays re-epithelialization; chronic wounds (diabetic ulcers) involve persistent inflammation and impaired migration.

Common Misconceptions and Exam Traps

• Wrong: "Intermediate filaments are static." Correct: They are more stable than actin or MTs but undergo regulated subunit exchange, and phosphorylation drives remodeling or disassembly at mitosis.
• Wrong: "All cytoskeletal filaments are polarized." Correct: IFs are apolar — tetramers assemble antiparallel, creating non-polarized filaments.
• Wrong: "Focal adhesions are just glue." Correct: They are signaling hubs — FAK, Src, and integrins transduce mechanical and chemical signals that regulate survival, proliferation, and migration.
• Wrong: "Rac and RhoA do the same thing." Correct: Rac promotes protrusion at the front; RhoA promotes contraction at the rear. Their antagonism is essential for polarized migration.