Dynamic aspects of the supramolecular organization of intermediate filament networks in cultured epidermal cells

Jonathan C.R. Jones*, Anne E. Goldman, Peter M. Steinert, Stuart Yuspa, Robert Goldman

*Corresponding author for this work

Research output: Contribution to journalArticle

70 Citations (Scopus)

Abstract

We have shown, by indirect immunofluorescence microscopy using an antiserum against the mouse keratin subunit K2 and by electron microscopy, that transformed (PAM) and primary (PME) mouse epidermal cells possess extensive net works of IF bundles. Following trypsinization and replating of PAM cells, IF bundles are seen to move as a continuous net work from a perinuclear zone into the peripheral cytoplasmic regions. In PAM cells lysed in high‐ionic‐strength solutions containing Triton ×‐100 and DNAase‐1, IF bundles appear to be closely associated with nuclear envelope remnants and, in some cases, appear to be attached to nuclear pore complexes. PME cells cultivated in low Ca2+‐containing medium possess perinuclear birefringent arrays of IF bundles. Within 2 hours of switching the cells to normal Ca2+ levels, the PME IF bundle network moves towards and establishes contact with the cell surface as desmosomes form. Live cells observed by phase contrast and fixed cells observed by immunofluorescence microscopy demonstrate that desmosomes can be distinguished as dark bands separating neighboring cells. There is little difference between the major proteins seen in SDS‐polyacrylamide gel profiles of isolated IF bundle net works from PME cells before and after the Ca2+ switch. Therefore, a reorganization of relatively insoluble membrane‐associated protein following the Ca2+ switch may be involved in desmosome formation. The isolated IF networks from PAM cells differ in protein composition compared to the PME IF networks. This may be related to the greatly reduced number of desmosomes in PAM cells. The IF bundle system in epidermal cells appears to be involved in shape formation, shape maintenance, the establishment of desmosomes, nuclear centration, and cell‐cell contact.

Original languageEnglish (US)
Pages (from-to)197-213
Number of pages17
JournalCell Motility
Volume2
Issue number3
DOIs
StatePublished - Jan 1 1982

Fingerprint

Intermediate Filaments
Cultured Cells
Desmosomes
Fluorescence Microscopy
Nuclear Pore
Proteins
Nuclear Envelope
Indirect Fluorescent Antibody Technique
Keratins
Immune Sera
Electron Microscopy

Keywords

  • desmosomes
  • epidermal keratinocytes
  • intermediate filament
  • nuclear envelope

ASJC Scopus subject areas

  • Cell Biology

Cite this

Jones, Jonathan C.R. ; Goldman, Anne E. ; Steinert, Peter M. ; Yuspa, Stuart ; Goldman, Robert. / Dynamic aspects of the supramolecular organization of intermediate filament networks in cultured epidermal cells. In: Cell Motility. 1982 ; Vol. 2, No. 3. pp. 197-213.
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Dynamic aspects of the supramolecular organization of intermediate filament networks in cultured epidermal cells. / Jones, Jonathan C.R.; Goldman, Anne E.; Steinert, Peter M.; Yuspa, Stuart; Goldman, Robert.

In: Cell Motility, Vol. 2, No. 3, 01.01.1982, p. 197-213.

Research output: Contribution to journalArticle

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N2 - We have shown, by indirect immunofluorescence microscopy using an antiserum against the mouse keratin subunit K2 and by electron microscopy, that transformed (PAM) and primary (PME) mouse epidermal cells possess extensive net works of IF bundles. Following trypsinization and replating of PAM cells, IF bundles are seen to move as a continuous net work from a perinuclear zone into the peripheral cytoplasmic regions. In PAM cells lysed in high‐ionic‐strength solutions containing Triton ×‐100 and DNAase‐1, IF bundles appear to be closely associated with nuclear envelope remnants and, in some cases, appear to be attached to nuclear pore complexes. PME cells cultivated in low Ca2+‐containing medium possess perinuclear birefringent arrays of IF bundles. Within 2 hours of switching the cells to normal Ca2+ levels, the PME IF bundle network moves towards and establishes contact with the cell surface as desmosomes form. Live cells observed by phase contrast and fixed cells observed by immunofluorescence microscopy demonstrate that desmosomes can be distinguished as dark bands separating neighboring cells. There is little difference between the major proteins seen in SDS‐polyacrylamide gel profiles of isolated IF bundle net works from PME cells before and after the Ca2+ switch. Therefore, a reorganization of relatively insoluble membrane‐associated protein following the Ca2+ switch may be involved in desmosome formation. The isolated IF networks from PAM cells differ in protein composition compared to the PME IF networks. This may be related to the greatly reduced number of desmosomes in PAM cells. The IF bundle system in epidermal cells appears to be involved in shape formation, shape maintenance, the establishment of desmosomes, nuclear centration, and cell‐cell contact.

AB - We have shown, by indirect immunofluorescence microscopy using an antiserum against the mouse keratin subunit K2 and by electron microscopy, that transformed (PAM) and primary (PME) mouse epidermal cells possess extensive net works of IF bundles. Following trypsinization and replating of PAM cells, IF bundles are seen to move as a continuous net work from a perinuclear zone into the peripheral cytoplasmic regions. In PAM cells lysed in high‐ionic‐strength solutions containing Triton ×‐100 and DNAase‐1, IF bundles appear to be closely associated with nuclear envelope remnants and, in some cases, appear to be attached to nuclear pore complexes. PME cells cultivated in low Ca2+‐containing medium possess perinuclear birefringent arrays of IF bundles. Within 2 hours of switching the cells to normal Ca2+ levels, the PME IF bundle network moves towards and establishes contact with the cell surface as desmosomes form. Live cells observed by phase contrast and fixed cells observed by immunofluorescence microscopy demonstrate that desmosomes can be distinguished as dark bands separating neighboring cells. There is little difference between the major proteins seen in SDS‐polyacrylamide gel profiles of isolated IF bundle net works from PME cells before and after the Ca2+ switch. Therefore, a reorganization of relatively insoluble membrane‐associated protein following the Ca2+ switch may be involved in desmosome formation. The isolated IF networks from PAM cells differ in protein composition compared to the PME IF networks. This may be related to the greatly reduced number of desmosomes in PAM cells. The IF bundle system in epidermal cells appears to be involved in shape formation, shape maintenance, the establishment of desmosomes, nuclear centration, and cell‐cell contact.

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