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Related Experiment Videos

Human epithelial cell intermediate filaments: isolation, purification, and characterization.

M W Aynardi, P M Steinert, R D Goldman

    The Journal of Cell Biology
    |April 1, 1984
    PubMed
    Summary
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    Human epithelial cell intermediate filaments (IF) can be disassembled and reassembled. Studies reveal HeLa IF share structural similarities with fibroblast IF proteins, indicating a common major IF protein in different cell types.

    Area of Science:

    • Cell Biology
    • Biochemistry
    • Structural Biology

    Background:

    • Intermediate filaments (IFs) are crucial cytoskeletal components.
    • Understanding the composition and assembly of IFs in different cell types is essential for cell biology.
    • HeLa cells, a human epithelial cell line, serve as a model for studying cellular structures.

    Purpose of the Study:

    • To characterize the protein composition of intermediate filaments (IF) from human epithelial (HeLa) cells.
    • To investigate the assembly properties and structural homology of HeLa IF with those from other cell types.
    • To identify distinct intermediate filament networks within HeLa cells.

    Main Methods:

    • Disassembly and reassembly of intermediate filaments (IF) in vitro.
    • Sodium dodecyl sulfate gel electrophoresis (SDS-PAGE) for protein analysis.

    Related Experiment Videos

  • Antibody production and immunofluorescence microscopy for cellular localization.
  • Amino acid composition, alpha-helix content, and peptide mapping for structural comparison.
  • Ultrastructural analysis of IF organization.

    • Main Results:

    • Eight proteins were identified in HeLa IF after disassembly-reassembly cycles.
    • Immunofluorescence revealed a complex pattern suggesting at least two distinct IF networks.
    • HeLa IF proteins share similarities in amino acid composition and alpha-helix content with other IF proteins.
    • The major 55,000-mol-wt HeLa IF protein shows homology with a similar protein from hamster fibroblasts (BHK-21).
    • In vitro reconstituted HeLa IF exhibited two organizational states: bundles and loose arrays.

    Conclusions:

    • The major intermediate filament (IF) protein in HeLa cells is structurally homologous to the major IF protein found in fibroblasts.
    • HeLa cells contain at least two distinct intermediate filament networks.
    • Intermediate filaments (IF) from human epithelial cells can be reconstituted in vitro, retaining distinct organizational states.