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Gel-expanded to gel-condensed transition in neurofilament networks revealed by direct force measurements

Nature Materials volume 9, pages 40–46 (2010)Cite this article

Abstract

Neurofilaments (NF)—the principal cytoskeletal constituent of myelinated axons in vertebrates—consist of three molecular-weight subunit proteins NF-L (low), NF-M (medium) and NF-H (high), assembled to form mature filaments with protruding unstructured C-terminus side arms1,2,3,4,5. Liquid-crystal gel networks of side-arm-mediated neurofilament assemblies have a key role in the mechanical stability of neuronal processes. Disruptions of the neurofilament network, owing to neurofilament over-accumulation or incorrect side-arm interactions, are a hallmark of motor-neuron diseases including amyotrophic lateral sclerosis3,4,5,6,7,8,9. Using synchrotron X-ray scattering, we report on a direct measurement of forces in reconstituted neurofilament gels under osmotic pressure (P). With increasing pressure near physiological salt and average phosphorylation conditions, NF-LMH, comprising the three subunits near in vivo composition, or NF-LH gels, undergo for P>Pc≈10 kPa, an abrupt non-reversible gel-expanded to gel-condensed transition. The transition indicates side-arm-mediated attractions between neurofilaments consistent with an electrostatic model of interpenetrating chains. In contrast, NF-LM gels remain in a collapsed state for P<Pc and transition to the gel-condensed state at P>Pc. These findings, which delineate the distinct roles of NF-M and NF-H in regulating neurofilament interactions, shed light on possible mechanisms for disruptions of optimal mechanical network properties.

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Figure 1: Hierarchical neurofilament self-assembly.
Figure 2: SAXS for different osmotic pressures and subunit compositions.
Figure 3: Osmotic pressure–distance curves for neurofilaments of different subunit protein compositions and buffer salinities.
Figure 4: Handshake analysis of complementary side-arm interactions.

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Acknowledgements

We thank R. Bruinsma, M. Kardar, Y. Li, A. Parsegian, P. Pincus, R. Podgornik, M. Wyrsta and E. Zhulina for discussion and correspondence and M. C. Choi for help with the AFM studies. The X-ray research was carried out at the Stanford Synchrotron Radiation Lightsource and at the Lawrence Berkeley National Laboratory Light Source. AFM experiments were conducted in the Microscopy & Microanalysis Facility of the UCSB Materials Research Laboratory (supported by NSF-MRSEC). C.R.S., J.D. and J.B.J. were supported by DOE BES DE-FG-02-06ER46314 (protein purification and assembly, characterization) and NSF DMR-0803103 (phase behaviour). R.B. was supported by the Human Frontier Science Program organization and in part by DOE BES.

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  1. and Developmental Biology Departments, Materials, Physics, and Molecular, Cellular, University of California Santa Barbara, California 93106, USA

    Roy Beck, Jayna B. Jones & Cyrus R. Safinya

  2. Chemistry and Biochemistry Department, University of California Santa Barbara, California 93106, USA

    Joanna Deek

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Contributions

This letter was written by R.B., C.R.S. and J.D.; R.B. collected and analysed the X-ray, AFM and microscopy data and assisted in purifying the protein; J.D. purified the proteins and collected the X-ray and microscopy data. J.B.J. carried out preliminary measurements and refined the protein purification protocol. C.R.S. designed the original study; R.B., J.D. and J.B.J. were involved in the study design. The data were interpreted by R.B., J.D. and C.R.S.; J.B.J. commented on the final manuscript.

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Correspondence to Roy Beck or Cyrus R. Safinya.

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Beck, R., Deek, J., Jones, J. et al. Gel-expanded to gel-condensed transition in neurofilament networks revealed by direct force measurements. Nature Mater 9, 40–46 (2010). https://doi.org/10.1038/nmat2566

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