Cytoskeleton - Intermediate Filaments

From CellBiology

ANAT3231 Cell Biology online lectures from the 2017 course.

Cytoskeleton - Intermediate Filaments

Introduction

Intermediate filament organization
IF keratin in keratinocytes (tem)

This lecture introduces the intermediate filaments, their structure and function within the cell. Unlike the microfilament and microtubule systems, the filaments themselves consist of a wide variety of different proteins. These intermediate filaments have important structural roles in cell integrity both internally and through specialized cellular junctions that occur between cell-cell and cell-matrix which surrounds them. This topic will be addressed again when we look at the cell cytoskeleton and the extracellular matrix.

Recent Keratin Review

Fanny Loschke, Kristin Seltmann, Jamal-Eddine Bouameur, Thomas M Magin Regulation of keratin network organization. Curr. Opin. Cell Biol.: 2015, 32;56-64 PubMed 25594948


"Keratins form the major intermediate filament cytoskeleton of epithelia and are assembled from heterodimers of 28 type I and 26 type II keratins in cell- and differentiation-dependent patterns. By virtue of their primary sequence composition, interactions with cell adhesion complexes and components of major signaling cascades, keratins act as targets and effectors of mechanical force and chemical signals to determine cell mechanics, epithelial cohesion and modulate signaling in keratin isotype-specific manners. Therefore, cell-specific keratin expression and organization impact on cell growth, migration and invasion. Here, we review the recent literature, focusing on the question how keratin networks are regulated and how the interplay of keratins with adhesion complexes affects these processes and provides a framework to understand keratins contribution to blistering and inflammatory disorders and to tumor metastasis."

Objectives

  • Broad understanding of the cell cytoskeleton
  • Understanding of intermediate filament function
  • Understanding of intermediate filament structure
  • Understanding of intermediate filament proteins
  • Understanding of intermediate filament location in nucleus and cytoplasm
  • Understanding of intermediate filament adhesion junctions
  • Broad understanding of intermediate filament abnormalities


Structure

Dimer structureIF vimentin cartoon desmin EM

Physical Characteristics

  • 10 nm diameter
  • Named by size relative to other cytoskeletal filaments
  • intermediate filaments have no structural polarity
  • Monomer - central α-helical domain
  • Dimer - 2 monomers form parallel coiled coil
  • Tetramer - pair of parallel dimers associates in an antiparallel staggered fashion
    • tetramer is the soluble subunit (analogous to MT αβ-tubulin dimer, or MF actin monomer)
  • Provide rope-like resistance to mechanical stress
  • In muscle- link Z discs of adjacent myofibrils
  • Organization can be altered by phosphorylation

IF Types

IF family Human keratin genes

  • Type I (n = 28)
    • Acidic keratins (pI < 5.7) 40–64 kDa
      • K9-28 (epithelia)
      • K31-40 (hair/nail)
  • Type II (n = 26)
    • Basic keratins (pI ≥ 6.0) 53–67 kDa
      • K1-8, K71-80 (epithelia)
      • K81-86 (hair/nail)

Keratins form heterodimers that assemble into heteropolymeric keratin filaments

  • Type III
    • Desmin (cardiac, skeletal and smooth muscle)
    • Vimentin (widespread distribution: leukocytes, blood vessels, endothelial, some epithelial and mesenchymal cells) 56 kDa
    • Peripherin (neurons) 57 kDa
    • Glial fibrillary acidic protein (GFAP) (astrocytes/glia) 50 kDa

Type III intermediate filament proteins can form both homo- and heteropolymeric filaments

  • Type IV
    • Neurofilament Low NF-L (neurons) 62 kDa
    • Neurofilament Medium NF-M (neurons) 110 kDa
    • Neurofilament High NF-H (neurons) 130 kDa

Neurofilaments form heteropolymers

    • α-internexin (CNS neurons)
    • Synemins (muscle)
    • Syncoilin (muscle)
    • Nestin (stem cell marker) 220 kDa
  • Type V
    • Lamin A/C (ubiquitous) 62–72 kDa
    • Lamin B1/2 (ubiquitous) 65–68 kDa
  • Orphan
    • Phakinin (lens)
    • Filensin (lens)

Intermediate Filament Associated Protein (IFAP)

  • Cross-link intermediate filaments with one another
    • forming a bundle (also called a tonofilament) or a network
  • IFAPs
    • Plectin 500 kDa Striated muscle, epithelia Nuclear envelop
    • Syncoilin 64 kDa Striated muscle
    • Nesprin-3 117 kDa Kerotinocytes
    • Paranemin 280 kDa
    • Desmuslin 230 kDa

Function

  • found in some metazoans (vertebrates, nematodes, and molluscs)
  • Desmin interacts with nebulin linking intermediate filament network and sarcomeres at the Z-discs
  • Keratin filament formation originates mainly from sites close to the actin-rich cell cortex
  • 2 alternate theories as to IF load/strain transmission
    • entropic gels - where no individual intermediate filaments experiences direct loading in tension
    • single intermediate filaments and bundles - extensible and elastic in vitro, and therefore well-suited to bearing tensional loads

Keratin Filaments and Desmosome Desmosome Muscle Desmin

Applications

Take advantage of the unique cell type expression pattern of IF proteins.

  • Nestin as a stem cell marker
  • GFAP can be used as an astrocyte marker in the analysis of neuronal tissue
  • Vimentin antibody can be used as a neural stem cell marker (ab45939)
  • Vimentin is highly expressed in fibroblasts and some expression in T- and B-lymphocytes. Expressed in many hormone-independent mammary carcinoma cell lines.

Abnormalities

Hutchinson-Gilford Progeria Syndrome
  • Cytoplasmic inclusions in several diseases Link
  • Hutchinson–Gilford progeria syndrome - Lamin A IF OMIM
  • Amyotrophic Lateral Sclerosis (ALS) - NF-H, peripherin OMIM
  • Alexander Disease - GFAP Genetics Home Reference OMIM
  • Charcot-Marie-Tooth disease - NF-L OMIM
  • Parkinson's Disease - NF-L, NF-M OMIM
  • Pachyonychia Congenita Genetics Home Reference
  • Giant Axonal Neuropathy Genetics Home Reference
  • Desminopathies desmin-related cardio-skeletal myopathy (CSM), also known as desmin-related myopathy (DRM)
    • 3 cardiomyopathy forms – dilated, hypertrophic and restrictive cardiomyopathies PMCID: 2592258

Inherited Epidermolysis Bullosa (EB) Medline Plus EB classification

  • Is characterized by a marked mechanical fragility of epithelial tissues and the formation of blisters, erosions and poorly healing skin ulcers.

Represents four major groups of skin diseases:

  • Epidermolysis Bullosa Simplex (EBS) - mutations in KRT14 and KRT5 genes Genetics Home Reference
  • Junctional epidermolysis bullosa (JEB) (gene products involved: laminin V, integrin, collagen, not IF related) GeneReviews
  • Dominant dystrophic epidermolysis bullosa (DDEB) (a collagen mutation not IF related)
  • Recessive dystrophic epidermolysis bullosa (RDEB) (a collagen mutation not IF related)

Links: Human Intermediate Filament Mutation Database | Genes and Diseases | OMIM - Online Mendelian Inheritance in Man

History

Below are some example historical research finding related to cell junctions from the JCB Archive.

1968 [1] [2] Not actin, not myosin, but intermediate. They are neither thick nor thin: Howard Holtzer identifies intermediate filaments as a completely new kind of filament.

References

Textbooks

Essential Cell Biology

  • Chapter 19 Tissues Epithelial sheets and Cell-Cell Junctions p606

Molecular Biology of the Cell

Alberts, Bruce; Johnson, Alexander; Lewis, Julian; Raff, Martin; Roberts, Keith; Walter, Peter New York and London: Garland Science; c2002


Molecular Cell Biology

Lodish, Harvey; Berk, Arnold; Zipursky, S. Lawrence; Matsudaira, Paul; Baltimore, David; Darnell, James E. New York: W. H. Freeman & Co.; c1999

The Cell- A Molecular Approach

Cooper, Geoffrey M. Sunderland (MA): Sinauer Associates, Inc.; c2000

Search Online Textbooks

IF Database

Books

PubMed

  • PubMed is a service of the U.S. National Library of Medicine that includes over 18 million citations from MEDLINE and other life science journals for biomedical articles back to 1948. PubMed includes links to full text articles and other related resources. PubMed
  • PubMed Central (PMC) is a free digital archive of biomedical and life sciences journal literature at the U.S. National Institutes of Health (NIH) in the National Library of Medicine (NLM) allowing all users free access to the material in PubMed Central. PMC
  • Online Mendelian Inheritance in Man (OMIM) is a comprehensive compendium of human genes and genetic phenotypes. The full-text, referenced overviews in OMIM contain information on all known mendelian disorders and over 12,000 genes. OMIM
  • Entrez is the integrated, text-based search and retrieval system used at NCBI for the major databases, including PubMed, Nucleotide and Protein Sequences, Protein Structures, Complete Genomes, Taxonomy, and others Entrez

Search Pubmed

Reviews

  • Intermediate filaments mediate cytoskeletal crosstalk. Chang L, Goldman RD. Nat Rev Mol Cell Biol. 2004 Aug;5(8):601-13. Review. PMID: 15366704
  • Intermediate filament scaffolds fulfill mechanical, organizational, and signaling functions in the cytoplasm. Kim S, Coulombe PA. Genes Dev. 2007 Jul 1;21(13):1581-97. Review. PMID: 17606637
  • The desmosome and pemphigus. Waschke J. Histochem Cell Biol. 2008 Jul;130(1):21-54. Epub 2008 Apr 3. Review. PMID: 18386043
  • Intermediate filaments: versatile building blocks of cell structure Goldman RD, Grin B, Mendez MG, Kuczmarski ER. Curr Opin Cell Biol. 2008 Feb;20(1):28-34. Epub 2008 Jan 4. Review. PMID: 18178072
  • Electron microscopy of intermediate filaments: teaming up with atomic force and confocal laser scanning microscopy. Kreplak L, Richter K, Aebi U, Herrmann H. Methods Cell Biol. 2008;88:273-97. Review. PMID: 18617039
  • Oshima RG. Intermediate filaments: a historical perspective. Exp Cell Res. 2007 Jun 10;313(10):1981-94. Epub 2007 Apr 11. Review. PMID: 17493611
  • Fuchs E, Cleveland DW. A structural scaffolding of intermediate filaments in health and disease. Science. 1998 Jan 23;279(5350):514-9. Review. PMID: 9438837

Articles

  • Visualization of a system of filaments 7-10 nm thick in cultured cells of an epithelioid line (Pt K2) by immunofluorescence microscopy. Osborn M, Franke WW, Weber K. Proc Natl Acad Sci U S A. 1977 Jun;74(6):2490-4. PMID: 329288
  • Different intermediate-sized filaments distinguished by immunofluorescence microscopy. Franke WW, Schmid E, Osborn M, Weber K. Proc Natl Acad Sci U S A. 1978 Oct;75(10):5034-8. PMID: 368806 PNAS

Acronyms

crescentin bacteria Caulobacter crescentus, with several properties similar to animal IF proteins

EBS epidermolysis bullosa simplex

filensin

GFAP glial gibrillary acidic protein

IF Intermediate Filament

IFAP Intermediate Filament Associated Protein

IFP Intermediate Filament Protein

KF Keratin filament

LMNA lamin A gene

NF neurofilament

NF-L neurofilament-low molecular mass protein

NF-M neurofilament-medium molecular mass protein

NF-H neurofilament-high molecular mass protein

phakinin

plectin an intermediate filament–bound cytolinker protein

vimentin Latin, vimentum for wickerwork or brushwood


Images



2015 Course Content

Lectures: Cell Biology Introduction | Cells Eukaryotes and Prokaryotes | Cell Membranes and Compartments | Cell Nucleus | Cell Export - Exocytosis | Cell Import - Endocytosis | Cytoskeleton Introduction | Cytoskeleton - Microfilaments | Cytoskeleton - Microtubules | Cytoskeleton - Intermediate Filaments | Cell Mitochondria | Cell Junctions | Extracellular Matrix 1 | Extracellular Matrix 2 | Cell Cycle | Cell Division | Cell Death 1 | Cell Death 2 | Signal 1 | Signal 2 | Stem Cells 1 | Stem Cells 2 | Development | 2015 Revision


Laboratories: Introduction to Lab | Microscopy Methods | Preparation/Fixation | Cell Knockout Methods | Cytoskeleton Exercise | Immunochemistry | Project Work | Confocal Microscopy | Tissue Culture | Stem Cells Lab | Microarray Visit

2015 Projects: Group 1 | Group 2 | Group 3 | Group 4 | Group 5 | Group 6 | Group 7

Dr Mark Hill 2015, UNSW Cell Biology - UNSW CRICOS Provider Code No. 00098G