Mechanical Effects of Neurofilament Cross-bridges

doi:10.1074/jbc.271.26.15687

Mechanical Effects of Neurofilament Cross-bridges

MODULATION BY PHOSPHORYLATION, LIPIDS, AND INTERACTIONS WITH F-ACTIN*

From ^‡ U.298 INSERM, CHRU, 49033 Angers Cedex, France
^§ Experimental Medicine Division, Brigham and Women's Hospital Boston Massachusetts 02115
^¶ Latvian Institute of Organic Synthesis, Aizkraukles Iela 21, Riga, LV-1006, Latvia, and
^| Program in Biological and Biomedical Sciences, Harvard Medical School, Boston, Massachusetts 02115

^” To whom correspondence should be addressed:
Experimental Medicine Division, Brigham and Women's Hospital, 221 Longwood Ave., Boston MA 02115.
Tel.: 617-278-0382; Fax: 617-734-2248; E-mail: janmey{at}fas.harvard.edu or janmey{at}calvin.bwh.harvard.edu.

Abstract

The structure of gels formed by bovine spinal cord neurofilaments was determined by fluorescence and electron microscopy and compared to mechanical properties measured by their elastic and viscous response to shear forces. Neurofilaments formed gels of high elastic modulus (>100 Pa) after addition of millimolar Mg²⁺. Gelation caused a slow increase in shear moduli to levels similar to those of vimentin intermediate filament networks, followed by a rapid rise due to formation of links between neurofilaments, mediated by cross-bridging structures that vimentin filaments lack. Neurofilament gels are more resistant to large deformations than are vimentin networks, suggesting the importance of cross-bridges for neurofilament mechanical properties.

Fluorescence imaging of single neurofilaments showed flexible filaments that became straighter when they adhered to glass or were incorporated into filament bundles. Electron microscopy of neurofilament gels showed a system of bundles intertwined within a more isotropic network of individual filaments.

Neurofilament gel formation was stimulated in vitro by acid phosphatase treatment or by inositol phospholipids. In contrast, addition of actin filaments reduced the resistance of neurofilament gels to large stresses. These results suggest that dynamic and regulated interactions occur between neurofilaments to form viscoelastic networks with properties distinct from other cytoskeletal structures.

Footnotes

↵* This work was supported in part by United States Public Health Service Grant AR38910, the Fogarty Foundation Grant TW00100, NATO Grant 940277, and European Community Grant INTAS 93–246. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

NF

neurofilament

MT

microtubule

Mes

4-morpholinepropanesulfonic acid

PIP₂

phosphatidylinositol 4,5-bisphosphate

IF

intermediate filament.
↵² J. F. Leterrier, unpublished observations.
↵³ A. Maggs and P. A. Janmey, unpublished data.
↵⁴ J. P. Gou and J. F. Leterrier, manuscript in preparation.
- Received December 26, 1995.
- Revision received April 5, 1996.