Rodolphe Perrot

Review of the Multiple Aspects of Neurofilament Functions, and their Possible Contribution to Neurodegeneration

Neurofilaments (NF) are the most abundant cytoskeletal component of large myelinated axons from adult central and peripheral nervous system. Here, we provide an overview of the complementary approaches, including biochemistry, cell biology and transgenic technology that were used to investigate the assembly, axonal transport and functions of NF in normal and pathological situations. Following their synthesis and assembly in the cell body, NFs are transported along the axon. This process is finely regulated via phosphorylation of the carboxy-terminal part of the two high-molecular-weight subunits of NF. The correct formation of an axonal network of NF is crucial for the establishment and maintenance of axonal calibre and consequently for the optimisation of conduction velocity. The frequent disorganisation of NF network observed in several neuropathologies support their contribution. However, despite the presence of NF mutations found in some patients, the exact relations between these mutations, the abnormal NF organisation and the pathological process remain a challenging field of investigation.

Neuronal intermediate filaments and neurodegenerative disorders

Intermediate filaments represent the most abundant cytoskeletal element in mature neurons. Mutations and/or accumulations of neuronal intermediate filament proteins are frequently observed in several human neurodegenerative disorders. Although it is now admitted that disorganization of the neurofilament network may be directly involved in neurodegeneration, certain type of perikaryal intermediate filament aggregates confer protection in motor neuron disease. The use of various mouse models provided a better knowledge of the role played by the disorganization of intermediate filaments in the pathogenesis of neurodegenerative disorders, but the mechanisms leading to the formation of these aggregates remain elusive. Here, we will review some neurodegenerative diseases involving intermediate filaments abnormalities and possible mechanisms susceptible to provoke them.

Reversal of neuropathy phenotypes in conditional mouse model of Charcot–Marie–Tooth disease type 2E

Mutations in the gene encoding for the neurofilament light subunit (NF-L) are responsible for Charcot-Marie-Tooth (CMT) neuropathy type 2E. To address whether CMT2E disease is potentially reversible, we generated a mouse model with conditional doxycycline-responsive gene system that allows repression of mutant hNF-LP22S transgene expression in adult neurons. The hNF-LP22S;tTa transgenic (tg) mice recapitulated key features of CMT2E disease, including aberrant hindlimb posture, motor deficits, hypertrophy of muscle fibres and loss of muscle innervation without neuronal loss. Remarkably, a 3-month treatment of hNF-LP22S;tTa mice with doxycycline after onset of disease efficiently down-regulated expression of hNF-LP22S and it caused reversal of CMT neurological phenotypes with restoration of muscle innervation and of neurofilament protein distribution along the sciatic nerve. These data suggest that therapeutic approaches aimed at abolishing expression or neutralizing hNF-L mutants might not only halt the progress of CMT2E disease, but also revert the disabilities.

Axonal Neurofilaments Control Multiple Fiber Properties But Do Not Influence Structure or Spacing of Nodes of Ranvier

In the vertebrate nervous system, axon calibers correlate positively with myelin sheath dimensions and electrophysiological parameters including action potential amplitude and conduction velocity. Neurofilaments, a prominent component of the neuronal cytoskeleton, are required by axons to support their normal radial growth. To distinguish between fiber features that arise in response to absolute axon caliber and those that are under autonomous control, we investigated transgenic mice in which neurofilaments are sequestered in neuronal cell bodies. The neurofilament deficient axons in such mice achieve mature calibers only 50% of normal and have altered conduction properties. We show here that this primary axonal defect also induces multiple changes in myelin sheath composition and radial dimensions. Remarkably, other fundamental fiber features, including internodal spacing and the architecture and composition of nodes of Ranvier, remain unaltered. Thus, many fiber characteristics are controlled through mechanisms operating independently of absolute axon caliber and the neurofilament cytoskeleton.

Real-time imaging reveals defects of fast axonal transport induced by disorganization of intermediate filaments

Intermediate filament (IF) abnormalities frequently appear in neurodegenerative disorders, but how they may contribute to neuronal dysfunction remains unclear. Here, we examined the effects of IF disorganization on the fast axonal transport using timelapse microscopy. We studied the axonal transport of mitochondria and lysosomes in cultured primary dorsal root ganglion (DRG) neurons derived from mice deficient for neurofilament light (NFL–/–), mice overexpressing peripherin (Per), and mice double transgenic Per;NFL–/–. Unexpectedly, a net retrograde transport of mitochondria was detected in Per;NFL–/– neurons, opposite to the net anterograde transport of these organelles observed in wild‐type (Wt), NFL–/–, and Per neurons. A detailed analysis of the kinetic properties of mitochondrial movements revealed an increased frequency of retrograde movements and an increase of their velocity in Per;NFL–/– neurons compared to Wt, NFL–/–, and Per neurons. We also noticed that the depletion of axonal neurofilaments (NFs) in NFL–/– and Per;NFL–/– neurons induced longer and more persistent movements of mitochondria and lysosomes in both directions, which suggests that the NF network hampers the traffic of these organelles. The finding that an up regulation of peripherin in context of NFL deficiency can provoke a net retrograde transport of mitochondria is a phenomenon that may contribute to pathogenic changes in some neurodegenerative disorders with IF protein accumulations.—Perrot, R., Julien, J.‐P. Realtime imaging reveals defects of fast axonal transport induced by disorganization of intermediate filaments. FASEB J. 23, 3213–3225 (2009). www.fasebj.org

Knockout Models of Neurofilament Proteins

Neurofilaments (NFs) are the most prominent cytoskeleton components of large myelinated axons from adult central and peripheral nervous systems. In the last 15 years, the gene targeting technique has been widely used to investigate the role of NF proteins in neuronal function. Gene knockout studies have demonstrated that NFs are crucial to expand the caliber of myelinated axons and consequently to increase their conduction velocity. However, the mechanism by which NFs determine the axonal diameter is not yet fully elucidated. NFs also contribute to the dynamic properties of the axonal cytoskeleton during neuronal differentiation, axon outgrowth and regeneration. Perturbations of their metabolism and organization are frequently associated with neurodegenerative disorders, including amyotrophic lateral sclerosis, Charcot-Marie-Tooth disease, Alzheimer’s disease and giant axonal neuropathy. Here, we describe how mouse knockout models of NF proteins have been used to study the multiple aspects of NF biology.

Crosstalks Between Myelinating Cells and the Axonal Cytoskeleton

A constant and dynamic communication between axons and myelinating cells is necessary for the correct development, function, and maintenance of myelinated fibers. Recently, several studies highlighted the pivotal role of the axonal cytoskeleton in this reciprocal communication. In particular, myelinating cells control the radial axonal growth by regulating the expression, transport, and organization of the axonal cytoskeleton. Conversely, this latter modulates dimensions of the myelin sheath by controlling the axonal caliber. Here, we will review the main investigations contributing to a better understanding of how the axoskeleton and myelinating cells influence each other to optimize conduction properties of myelinated fibers.

Maldistribution of Neurofilaments, Disease Pathogenesis, and Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS), also called Lou Gehrig’s disease, is a late-onset progressive motor neuron disease first identified in 1869 by Jean-Martin Charcot. Its incidence is approximately two cases per 100,000, with a slightly higher prevalence in men. The progressive degeneration of motor neurons in ALS leads to neuron death and loss of muscle function. Patients become partially or totally paralyzed, but their cognitive functions usually remain unaffected. There is no cure for ALS and the disease is usually fatal within three to five years of the onset of symptoms. About 90% of ALS cases are sporadic, while approximately 10% are inherited in a dominant manner. Mutations in the gene coding for superoxide dismutase 1 (SOD1) account for 20% of all familial cases. To date, more than 140 mutations have been found in the SOD1 gene, and transgenic mice overexpressing various SOD1 mutants develop an ALS-like phenotype through a gain of unknown toxic properties.