Disruption of neurofilament network with aggregation of light neurofilament protein: a common pathway leading to motor neuron degeneration due to Charcot-Marie-Tooth disease-linked mutations in NFL and HSPB1. Hum Mol Genet

Division of Neuropathology, Department of Pathology and Laboratory Medicine, School of Medicine, University of Pennsylvania, Philadelphia, PA 19104,-6100, USA.
Human Molecular Genetics (Impact Factor: 6.39). 01/2008; 16(24):3103-16. DOI: 10.1093/hmg/ddm272
Source: PubMed


Mutations in neurofilament light (NFL) subunit and small heat-shock protein B1 (HSPB1) cause autosomal-dominant axonal Charcot-Marie-Tooth disease type 2E (CMT2E) and type 2F (CMT2F). Previous studies have shown that CMT mutations in NFL and HSPB1 disrupt NF assembly and cause aggregation of NFL protein. In this study, we investigate the role of aggregation of NFL protein in the neurotoxicity of CMT mutant NFL and CMT mutant HSPB1 in motor neurons. We find that expression of CMT mutant NFL leads to progressive degeneration and loss of neuronal viability of cultured motor neurons. Degenerating motor neurons show fragmentation and loss of neuritic processes associated with disruption of NF network and aggregation of NFL protein. Co-expression of wild-type HSPB1 diminishes aggregation of CMT mutant NFL, induces reversal of CMT mutant NFL aggregates and reduces CMT mutant NFL-induced loss of motor neuron viability. Like CMT mutant NFL, expression of S135F CMT mutant HSPB1 also leads to progressive degeneration of motor neurons with disruption of NF network and aggregation of NFL protein. Further studies show that wild-type and S135F mutant HSPB1 associate with wild-type and CMT mutant NFL and that S135F mutant HSPB1 has dominant effect on disruption of NF assembly and aggregation of NFL protein. Finally, we show that deletion of NFL markedly reduces degeneration and loss of motor neuron viability induced by S135F mutant HSPB1. Together, our data support the view that disruption of NF network with aggregation of NFL is a common triggering event of motor neuron degeneration in CMT2E and CMT2F disease.

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Available from: Hong Lin, Dec 18, 2013
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    • "Several studies have found cytoskeletal abnormalities and axonal transport defects. For instance, overexpression of HSPB1 S135F or HSPB1 P182L led to aggregation and altered axonal transport of neurofilament [6] [31] [32]. These mutants also displayed increased binding to tubulin, which led to stabilization and altered dynamics of microtubules [33]. "
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    ABSTRACT: Background: HSPB1 belongs to the family of small heat shock proteins (sHSP) that have importance in protection against unfolded protein stress, in cancer cells for escaping drug toxicity stress and in neurons for suppression of protein aggregates. sHSPs have a conserved α-crystalline domain (ACD), flanked by variable N- and C-termini, whose functions are not fully understood. Dominant missense variants in HSPB1, locating mostly to the ACD, have been linked to inherited neuropathy. Methods: Patients underwent detailed clinical and neurophysiologic characterization. Disease causing variants were identified by exome or gene panel sequencing. Primary patient fibroblasts were used to investigate the effects of the dominant defective HSPB1 proteins. Results: Frameshift variant predicting ablation of the entire C-terminus p.(Met169Cfs2*) of HSPB1 and a missense variant p.(Arg127Leu) were identified in patients with dominantly inherited motor-predominant axonal Charcot-Marie-Tooth neuropathy. We show that the truncated protein is stable and binds wild type HSPB1. Both mutations impaired the heat stress tolerance of the fibroblasts. This effect was particularly pronounced for the cells with the truncating variant, independent of heat-induced nuclear translocation and induction of global transcriptional heat response. Furthermore, the truncated HSPB1 increased cellular sensitivity to protein misfolding. Conclusion: Our results suggest that truncation of the non-conserved C-terminus impairs the function of HSPB1 in cellular stress response. General significance: sHSPs have important roles in prevention of protein aggregates that induce toxicity. We showed that C-terminal part of HSPB1 is critical for tolerance of unfolded protein stress, and when lacking causes axonal neuropathy in patients.
    Full-text · Article · Mar 2015 · Biochimica et Biophysica Acta - Clinical
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    • "However, the pleiotropic nature of the HSPB1 protein suggests that some neuron-specific functions could contribute to peripheral neuropathy. Indeed, CMT-causing mutations in HSPB1 have been shown to cause aggregation of the neurofilament light protein, and therefore disruption of the neurofilament (NF) network [26, 74]. "
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    ABSTRACT: Mutations in the small heat shock protein HSPB1 (HSP27) are a cause of axonal Charcot–Marie–Tooth neuropathy (CMT2F) and distal hereditary motor neuropathy. To better understand the effect of mutations in HSPB1 on the neuronal cytoskeleton, we stably transduced neuronal cells with wild-type and mutant HSPB1 and investigated axonal transport of neurofilaments (NFs). We observed that mutant HSPB1 affected the binding of NFs to the anterograde motor protein kinesin, reducing anterograde transport of NFs. These deficits were associated with an increased phosphorylation of NFs and cyclin-dependent kinase Cdk5. As Cdk5 mediates NF phosphorylation, inhibition of Cdk5/p35 restored NF phosphorylation level, as well as NF binding to kinesin in mutant HSPB1 neuronal cells. Altogether, we demonstrate that HSPB1 mutations induce hyperphosphorylation of NFs through Cdk5 and reduce anterograde transport of NFs. Electronic supplementary material The online version of this article (doi:10.1007/s00401-013-1133-6) contains supplementary material, which is available to authorized users.
    Full-text · Article · Jun 2013 · Acta Neuropathologica
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    • "There have been many suggestions to explain the tissue-specific pathologies associated with sHSP mutations [34–40], but we propose that the specific intermediate filament expression pattern must be considered as a key factor in any sHSP-based pathology as we consider the intermediate filament–sHSP complex to be a functional unit [41,42]. Intermediate filament expression profiles follow tissue-specific patterns according to embryological origins [43]. "
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    ABSTRACT: CRYAB (αB-crystallin) is expressed in many tissues and yet the R120G mutation in CRYAB causes tissue-specific pathologies, namely cardiomyopathy and cataract. Here, we present evidence to demonstrate that there is a specific functional interaction of CRYAB with desmin intermediate filaments that predisposes myocytes to disease caused by the R120G mutation. We use a variety of biochemical and biophysical techniques to show that plant, animal and ascidian small heat-shock proteins (sHSPs) can interact with intermediate filaments. Nevertheless, the mutation R120G in CRYAB does specifically change that interaction when compared with equivalent substitutions in HSP27 (R140G) and into the Caenorhabditis elegans HSP16.2 (R95G). By transient transfection, we show that R120G CRYAB specifically promotes intermediate filament aggregation in MCF7 cells. The transient transfection of R120G CRYAB alone has no significant effect upon cell viability, although bundling of the endogenous intermediate filament network occurs and the mitochondria are concentrated into the perinuclear region. The combination of R120G CRYAB co-transfected with wild-type desmin, however, causes a significant reduction in cell viability. Therefore, we suggest that while there is an innate ability of sHSPs to interact with and to bind to intermediate filaments, it is the specific combination of desmin and CRYAB that compromises cell viability and this is potentially the key to the muscle pathology caused by the R120G CRYAB.
    Full-text · Article · May 2013 · Philosophical Transactions of The Royal Society B Biological Sciences
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