The overall goal of this proposal is to understand the structure and function of the neuronal cytoskeleton. The proposed studies are designed to give us a basic knowledge of the function of the neuronal cytoskeleton in the normal CNS. Towards this end, we will use a wide variety of techniques to study the expression, assembly, phosphorylation and interactions of a number of neuronal cytoskeletal proteins. Ultimately, we hope that this information will help us understand why and how the neuronal cytoskeleton is perturbed by certain neurological diseases, such as Alzheimer's disease and amyotropic lateral sclerosis and by exposure to neurotoxins, such as aluminum and acrylamide.
The specific aims of this proposal are 1) to study the interactions and assembly of the three neurofilament proteins using purified enzymatic and cyanogen bromide derived fragments of each neurofilament triplet molecule to determine the role of phosphorylation of these three proteins in the neurofilament assembly; 2) to identify and isolate the kinase and/or phosphatase responsible for the phosphorylation of the neurofilament proteins; 3) to continue our studies on the interactions of neurofilaments and microtubules, in order to define which microtubule associated proteins (MAPs) are involved in these interactions; 4) to study the importance of the cytoskeletal associated ATPase, Beta-internexin in cytoskeleton-mediated cytoplasmic transport in neurons and cultured cells and 5) to obtain cDNA probes for the high molecular weight neurofilament proteins, NF-M and NF-H, which will be used to study the expression and function of these proteins and to study the assembly of various modified proteins. The cDNA clones will be placed in expression vectors and their distribution will be studied in cells, which do not normally contain neurofilaments. These studies will allow us to determine what structures they form in the cell and what their function might be.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS015182-10
Application #
3396022
Study Section
Neurology B Subcommittee 1 (NEUB)
Project Start
1987-07-01
Project End
1993-06-30
Budget Start
1988-07-01
Budget End
1989-06-30
Support Year
10
Fiscal Year
1988
Total Cost
Indirect Cost
Name
Columbia University (N.Y.)
Department
Type
Schools of Medicine
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10027
Liem, Ronald K H; Messing, Albee (2009) Dysfunctions of neuronal and glial intermediate filaments in disease. J Clin Invest 119:1814-24
Ching, Gee Y; Liem, Ronald K H (2009) RE1 silencing transcription factor is involved in regulating neuron-specific expression of alpha-internexin and neurofilament genes. J Neurochem 109:1610-23
Goryunov, Dmitry; Nightingale, Andrew; Bornfleth, Lorelei et al. (2008) Multiple disease-linked myotubularin mutations cause NFL assembly defects in cultured cells and disrupt myotubularin dimerization. J Neurochem 104:1536-52
Kabzinska, Dagmara; Perez-Olle, Raul; Goryunov, Dmitry et al. (2006) Is a novel I214M substitution in the NEFL gene a cause of Charcot-Marie-Tooth disease? Functional analysis using cell culture models. J Peripher Nerv Syst 11:225-31
Perez-Olle, Raul; Lopez-Toledano, Miguel A; Goryunov, Dmitry et al. (2005) Mutations in the neurofilament light gene linked to Charcot-Marie-Tooth disease cause defects in transport. J Neurochem 93:861-74
Perez-Olle, Raul; Jones, Sidonie T; Liem, Ronald K H (2004) Phenotypic analysis of neurofilament light gene mutations linked to Charcot-Marie-Tooth disease in cell culture models. Hum Mol Genet 13:2207-20
Perez-Olle, Raul; Lopez-Toledano, Miguel A; Liem, Ronald K H (2004) The G336S variant in the human neurofilament-M gene does not affect its assembly or distribution: importance of the functional analysis of neurofilament variants. J Neuropathol Exp Neurol 63:759-74
Leung, Conrad L; He, Cui Zhen; Kaufmann, Petra et al. (2004) A pathogenic peripherin gene mutation in a patient with amyotrophic lateral sclerosis. Brain Pathol 14:290-6
Leung, Conrad L; Green, Kathleen J; Liem, Ronald K H (2002) Plakins: a family of versatile cytolinker proteins. Trends Cell Biol 12:37-45
Perez-Olle, Raul; Leung, Conrad L; Liem, Ronald K H (2002) Effects of Charcot-Marie-Tooth-linked mutations of the neurofilament light subunit on intermediate filament formation. J Cell Sci 115:4937-46

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