Neurofilaments (NF), the major cytoskeletal proteins expressed in mature myelinated axons, are intrinsic determinants of axonal diameter (caliber). Maintenance of axonal caliber is important since conduction velocity of nerve action potentials is a function of axonal diameter. NF modulation of axonal caliber appears to be determined not by NF number, per se, but by their phosphorylation state. The mechanisms controlling NF phosphorylation are not understood. However, abnormal phosphorylation and accumulation of NF proteins are features of Alzheimer's disease (AD) and the motor neuron disease amyotrophic lateral sclerosis (ALS). The applicant is studying protein kinases that phosphorylate NF using mouse as a model system. He has identified and characterized a novel 115 kDa NF-associated kinase from mouse brain extracts, which has been named NAK115. NAK115 is expressed in neurons and its activity is down-regulated during myelination induced in culture. A kinase related to NAK115 is found in human brain. In addition to NAK115 two other NF kinases have been discovered. One of these appears to be a novel cdc2-like kinase that is associated with NF in mouse brain. A kinases with similar immunological properties has also been found in humans and is associated with the paired helical filaments that comprise the neurofibrillary tangles of Alzheimer disease. A 35 kDa NF kinase whose activity is up-regulated by myelination has also been discovered. In order to fully understand the regulation NF phosphorylation by these kinases, the applicant proposes to characterize the kinases biochemically, immunologically and at the molecular level. In a consortium study, it will also be determined whether the three NF kinases are associated with NF proteins during axonal transport in nerve fibers. In these studies it will also be determined whether axonal transport of the kinases are impaired to the same extent as the NF proteins after systemic intoxication with IDPN (beta,beta'-iminodipropionitrile), a neurotoxin that selectively blocks NF transport without significantly altering the transport of other cytoskeletal proteins, such as actin or tubulin. Finally, the applicant proposes to study how expression of the NF kinases changes during peripheral nerve injury when the cytoskeleton is restructured. These studies should provide valuable information on the association and transport of kinases with NF proteins, and role of the kinases in modification of NF proteins in neuronal function during normal development, during nerve injury, and in human diseases in which NF are affected.
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