Fig. 1

NFs structure, physiological processes and pathological modifications in ALS. The cartoon’s upper panel shows the physiological structure and composition of NF isoforms; note the common structure with a variable N-terminal head and the conserved central coiled coil across all isoforms on the left window. The variably elongated C-terminal tail domain, particularly represented in NfM and NfH, is rich in KSP repeats, which are sites for post-translational phosphorylation. Main post-translational modifications (central window), include glycosylation through the attachment of O-linked N-acetylglucosamine (O-GlcNAc) to individual Ser and Thr residues by O-linked N-acetylglucosamine transferase (OGT) and phosphorylation, which are reciprocal processes, i.e., reduced glycosylation may result in excess harmful phosphorylation [41]. Phosphorylation results from the tightly regulated activity of several kinases and phosphatases commanded by second-messenger pathways. The head domain of every NF isoform is phosphorylated in the cell body by protein kinase A (PKA), protein kinase C (PKC), calcium/calmodulin-dependent protein kinase type II (CAMKII) and CDK5 [24]. In contrast, phosphorylation in the tail KSP repeats almost exclusively occurs in NfM, and NfH on their entry into and transit along the axon. This last process is highly dynamic and seems to be involved in the axonal transport of NF during myelination or synaptogenesis; however, it may also be triggered by stress phenomena. The pairing of parallel heterodimers and antiparallel tetramers, by interactions between coil domains, mediates the assembly of NF isoforms; eight tetramers associate to form cylindrical structures known as unit-length filaments (ULFs), which approximate the final diameter of NF. End-to-end annealing of ULFs allows longitudinal elongation, while radial compaction results in the final width of 10 um in the mature NF polymer. The net forward movement of NF along the axons (right upper window) is the result of a rapid intermittent phase driven by molecular motors (dynein and dynactin) carrying hetero-oligomers along the microtubule and pauses which are determined by reversible attachment of NFs. However, the majority of NF isoforms resides in a stable network, representing the stationary phase of NF transport along the axon [24]. Molecular aberrations within these physiological processes may result in motor neuron pathologies, as shown in the lower panel. In the first window on the left, red triangles highlight the mutations in NF isoforms known to cause ALS. Modifications in subunit stoichiometry by overexpression of one isoform over the others may result in pathological aggregates. For example, overexpression of peripherin or mutated NfL can cause ALS-like neurofilament aggregates and selective degeneration of spinal motor neurons, and NfM and NfH dysregulated upregulation, often accompanied by excessive phosphorylation, may result in aberrant perikaryal neurofilament aggregates [42]. Excessive phosphorylation may be promoted by reduced glycosylation activity due to a loss-of-function mutation in a glycosyltransferase, GLT8D1 [43]. Glutamate excitotoxicity activates several kinases (MAPK,PKN11, PIN1) which end up in NfM and NfH tail hyperphophorylation. This last process has been repeatedly associated with two major histopathological findings in ALS, namely hyaline conglomerate inclusions (HCIs), ubiquitin-negative floccular inclusions rich in neurofilaments and peripherin, and axonal spheroids, which are non-pathognomonic for ALS but seems to occur early during motor neuron degeneration in fALS and sALS (right lower window). Studies on hyperphosphorylation of NfM and NfH showed that this phenomenon may lead to a slowing of NF transport along the axon, with accumulation of Nf inclusions outside the nucleus and in the axon, engorgement of perikaryal structures and disrupted dynamics of axonal circuitry [44]. Abbreviations: NfL, neurofilament light chain; NfM, neurofilament medium chain; NfH, neurofilament heavy chain; E segment, glutamic-acid-rich segment; E1, glutamic-acid rich segment 1; E2, glutamic-acid-rich segment 2; KE, lysine–glutamic acid; KEP, lysine–glutamic acid–proline; SP, serine–proline; KSP, lysine–serine–proline; O-GlcNAc, O-linked N-acetylglucosamine; OGT, intracellular glycosyltransferase O-linked N-acetylglucosamine transferase; EOGT, extracellular glycosyltransferase EGF domain-specific O-linked N-acetylglucosamine transferase; OGA, O-GlcNAcase; PKA, protein kinase A; PKC, protein kinase C; CAMKII, calcium/calmodulin-dependent protein kinase type II; CDK5, cyclin-dependent kinase-5; MAPK, mitogen-activated protein kinase; ERK, extracellular signal-regulated kinase; JNK, c-Jun N-terminal kinases; GSK-3, Glycogen synthase kinase 3;GLT8D1, glycosyltransferase 8 domain-containing 1; PKN11, Serine/threonine protein kinase C-related kinase; PIN1, Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1