Abstract
The degradation of phosphorylated and dephosphorylated neurofilament proteins by the Ca2+-activated neutral proteinase calpain was studied. Neurofilaments were isolated from bovine spinal cord, dephosphorylated by alkaline phosphatase (from Escherichia coli) and radioiodinated with [125I]-Bolton-Hunter reagent. The radioiodinated neurofilament proteins (untreated and dephosphorylated) were incubated in the presence and absence of calpain from rabbit skeletal muscle, and the degradation rates of large (NF-H), mid-sized (NF-M) and small (NF-L) neurofilament polypeptides were analysed by SDS/polyacrylamide-gel electrophoresis and autoradiography. The degradation of dephosphorylated neurofilament proteins occurred at a higher rate, and to a greater extent, than did that of the phosphorylated (untreated) neurofilament proteins. The dephosphorylated high-molecular-mass neurofilament (NF-HD) was proteolyzed 6 times more quickly than the untreated NF-H. The degradation rate of the NF-M and NF-L neurofilament proteins was also enhanced after dephosphorylation, but less than that of NF-H. This indicates that the dephosphorylation of neurofilament proteins can increase their sensitivity to calpain degradation.
Full text
PDF



Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bass M., Pant H. C., Gainer H., Soderling T. R. Calcium/calmodulin-dependent protein kinase II in squid synaptosomes. J Neurochem. 1987 Oct;49(4):1116–1123. doi: 10.1111/j.1471-4159.1987.tb10001.x. [DOI] [PubMed] [Google Scholar]
- Bennett V. Proteins involved in membrane--cytoskeleton association in human erythrocytes: spectrin, ankyrin, and band 3. Methods Enzymol. 1983;96:313–324. doi: 10.1016/s0076-6879(83)96029-9. [DOI] [PubMed] [Google Scholar]
- Black M. M., Keyser P., Sobel E. Interval between the synthesis and assembly of cytoskeletal proteins in cultured neurons. J Neurosci. 1986 Apr;6(4):1004–1012. doi: 10.1523/JNEUROSCI.06-04-01004.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Brown A., Eagles P. A. Squid neurofilaments. Phosphorylation and Ca2+-dependent proteolysis in situ. Biochem J. 1986 Oct 1;239(1):191–197. doi: 10.1042/bj2390191. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cantino D., Mugnaini E. The structural basis for electrotonic coupling in the avian ciliary ganglion. A study with thin sectioning and freeze-fracturing. J Neurocytol. 1975 Oct;4(5):505–536. doi: 10.1007/BF01351535. [DOI] [PubMed] [Google Scholar]
- Carden M. J., Schlaepfer W. W., Lee V. M. The structure, biochemical properties, and immunogenicity of neurofilament peripheral regions are determined by phosphorylation state. J Biol Chem. 1985 Aug 15;260(17):9805–9817. [PubMed] [Google Scholar]
- Cohen R. S., Pant H. C., House S., Gainer H. Biochemical and immunocytochemical characterization and distribution of phosphorylated and nonphosphorylated subunits of neurofilaments in squid giant axon and stellate ganglion. J Neurosci. 1987 Jul;7(7):2056–2074. doi: 10.1523/JNEUROSCI.07-07-02056.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Georges E., Lefebvre S., Mushynski W. E. Dephosphorylation of neurofilaments by exogenous phosphatases has no effect on reassembly of subunits. J Neurochem. 1986 Aug;47(2):477–483. doi: 10.1111/j.1471-4159.1986.tb04526.x. [DOI] [PubMed] [Google Scholar]
- Goldstein M. E., Sternberger L. A., Sternberger N. H. Microheterogeneity ("neurotypy") of neurofilament proteins. Proc Natl Acad Sci U S A. 1983 May;80(10):3101–3105. doi: 10.1073/pnas.80.10.3101. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goldstein M. E., Sternberger N. H., Sternberger L. A. Phosphorylation protects neurofilaments against proteolysis. J Neuroimmunol. 1987 Mar;14(2):149–160. doi: 10.1016/0165-5728(87)90049-x. [DOI] [PubMed] [Google Scholar]
- Julien J. P., Smoluk G. D., Mushynski W. E. Characteristics of the protein kinase activity associated with rat neurofilament preparations. Biochim Biophys Acta. 1983 Jan 4;755(1):25–31. doi: 10.1016/0304-4165(83)90268-4. [DOI] [PubMed] [Google Scholar]
- Lee V. M., Carden M. J., Trojanowski J. Q. Novel monoclonal antibodies provide evidence for the in situ existence of a nonphosphorylated form of the largest neurofilament subunit. J Neurosci. 1986 Mar;6(3):850–858. doi: 10.1523/JNEUROSCI.06-03-00850.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Neville D. M., Jr Molecular weight determination of protein-dodecyl sulfate complexes by gel electrophoresis in a discontinuous buffer system. J Biol Chem. 1971 Oct 25;246(20):6328–6334. [PubMed] [Google Scholar]
- Oblinger M. M. Characterization of posttranslational processing of the mammalian high-molecular-weight neurofilament protein in vivo. J Neurosci. 1987 Aug;7(8):2510–2521. [PMC free article] [PubMed] [Google Scholar]
- Paggi P., Lasek R. J. Axonal transport of cytoskeletal proteins in oculomotor axons and their residence times in the axon terminals. J Neurosci. 1987 Aug;7(8):2397–2411. [PMC free article] [PubMed] [Google Scholar]
- Pant H. C., Gallant P. E., Gainer H. Characterization of a cyclic nucleotide- and calcium-independent neurofilament protein kinase activity in axoplasm from the squid giant axon. J Biol Chem. 1986 Feb 25;261(6):2968–2977. [PubMed] [Google Scholar]
- Pant H. C., Gallant P. E., Gould R., Gainer H. Distribution of calcium-activated protease activity and endogenous substrates in the squid nervous system. J Neurosci. 1982 Nov;2(11):1578–1587. doi: 10.1523/JNEUROSCI.02-11-01578.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pant H. C., Terakawa S., Baumgold J., Tasaki I., Gainer H. Protein release from the internal surface of the squid giant axon membrane during excitation and potassium depolarization. Biochim Biophys Acta. 1978 Oct 19;513(1):132–140. doi: 10.1016/0005-2736(78)90118-9. [DOI] [PubMed] [Google Scholar]
- Sternberger L. A., Sternberger N. H. Monoclonal antibodies distinguish phosphorylated and nonphosphorylated forms of neurofilaments in situ. Proc Natl Acad Sci U S A. 1983 Oct;80(19):6126–6130. doi: 10.1073/pnas.80.19.6126. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Trojanowski J. Q., Walkenstein N., Lee V. M. Expression of neurofilament subunits in neurons of the central and peripheral nervous system: an immunohistochemical study with monoclonal antibodies. J Neurosci. 1986 Mar;6(3):650–660. doi: 10.1523/JNEUROSCI.06-03-00650.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]


