Molecular Cell Biology Intermediate Filaments Cooper

Introduction n Filaments 10 nm wide => “intermediate” n Present in Metazoa / Animals i.e. not Plants or Unicellular Organisms n Complex Gene Superfamily 70 in Human Genome n Specific Expression at Different Times and Places

Intermediate Filament Biochemical Properties In Vitro n Very stable. Little subunit exchange. n Very strong. Filaments do not break. MT’s strong but brittle Actin weak

Intermediate Filament Potential Functions In Vivo n Mechanical Strength of Cytoplasm n Help a Layer of Epithelial Cells Resist Shear Stress - Filaments Connect to Cell-cell Junctions n Hold Nucleus in Center of Cell

Intermediate Filament Structure & Assembly

Intermediate Filaments by EM: Filament Unraveling

Classes of Intermediate Filaments

Regulation of IF Assembly n Notoriously Stable No Nucleotide n Filaments Move Little Precursors Move More n Disassemble Somewhat during Mitosis Phosphorylation by Cyclin-depen Kinase

Vimentin Filaments in a Cultured Cell

Vimentin n All Cells in Early Development n Cage Around Nucleus n Interacts with Mt’s n Vimentin Knockout Mouse Initially normal at gross inspection Cultured cells have altered properties of uncertain significance

FRAP of Vimentin vs. Keratin in One Cell Left: Vimentin (Green) Right: Keratin (Red) 10 min time intervals

Dynamics of Keratin Particles in Periphery 11 micrometers over 10 minutes 18 micrometers over 10 minutes

Desmin n Expressed in Muscle n Elastic Elements to Prevent Over-stretching n Connects / Aligns Z lines n Knockout Mouse - Deranged Myofibril Architecture

Keratins n Expressed in Epithelia n Keratin Filaments Connect to Desmosome and Hemidesmosomes n Differentiation of Epidermis includes Production of Massive Amounts of Keratin n Provides Outer Protection of Skin n Composes Hair, Nails, Feathers, etc.

Density of Keratin Filaments in Outer Epidermis Layers

Keratin Mutations are Basis for Human Epidermal Diseases n Structure/Function Analysis of Keratin Assembly n Point Mutation in Terminal Domain Fails to Assemble n Mutant is Dominant, even in Low Amounts, in Cultured Cells and Mice

Epidermolysis Bullosa Simplex Wild-typeMutant

Keratins and EBS

Neurons n Neurofilament H, M, L Copolymer n Prevent Axon Breakage n Diseases with Clumps of Neurofilaments Superoxide dismutase model for ALS Clumps are secondary, not causative

Neurofilament Transport in Axons Photobleached Zone in the Middle

Neurofilament Transport in Axons Photobleached Zone in the Middle

Lamins n Square Lattice on Inner Surface of Nuclear Membrane n Present in Metazoans (Animals, not Plants or unicellular organisms) n Mitosis Breakdown Phosphorylation of A & C by Cyclin-depen Kinase B remains with Membrane n Mutations Cause Accelerated Aging Diseases Progerias - Dominant Mutations

EM of Nuclear Lamina Nuclear Pores

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