Cytoskeletons and cell death-associated proteins expressed in neuronal cells in the neurofilament gene-manipulated mice

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Neurofilaments (NFs) are composed of three proteins, NF-L,NF-M and NP-H, and NF-M and NP-H are provided with uniquely extended and phosphorylated carboxyl-terminal tail domains that do not appear in other intermediate filament proteins. In collaboration with American researchers at California (UCSD) and New York Universities we generated knockout mice that do not express each of these subunit proteins as well as knockin mice that lack only the carboxyl-terminal tail of NF-M or/and NF-H, to know how the deletions of each NF subunit protein and of the extended carboxy-terminal tails of NF-M or/and NF-H are related with NF organization in the neuron, axonal structure, axonal transport of NFs, organizations of other cytoskeletons and membrane-bound organelles, and expression of other NF subunit proteins. When NF-L was deleted, NFs were not assembled in neurons. Such axons were much thinner but provided with many more microtubules and mitochondria. Compared with NF-H, the deletion of NF-M had more effective influence on axonal calibers and NF organization, suggesting that NF-M is more essential for the NF function. Since the carboxyl-terminal tail of NF-M appears to be more important for NF organization and function, we deleted only this tail domain of NF-M or/and NF-H, and found as expected that NF-M tail was more necessary for the NF organization. However, more drastic changes in axoplasmic organizations, such as irregularity of NF alignment, Increase in density of microtubules as well as mitochondria, were obtained when both NF-M and NF-H tails were simultaneously deleted. These results suggest that both NF-M and NP-H tails, which form crossbridges between NFs, are necessary for NF organization and axonal functions, and are also associated with regulation of neuronal cell death through the change in function of mitochondria.