Cloning of m-calpain 80 kD subunit from the axonal degeneration-resistant WLD(S) mouse mutant.
Calpains are calcium-activated cysteine proteases that are involved in cellular degradation in models of neurodegeneration. Calpains are the effectors of cytoskeletal disruption during axonal degeneration, a pathological feature of many neurological disorders. The WLD(S) mouse mutant is resistant to axonal degeneration and demonstrates prolonged survival of the cytoskeleton after nerve injury. To investigate the possibility that a mutation in calpain or abnormalities in calpain protein expression is responsible for the resistance to axonal degeneration seen in the WLD(S) mouse mutant, we 1) cloned and sequenced the large subunit of the high calcium-requiring form of calpain (m-calpain) from nervous system tissues of WLD(S) and from wild-type C57BL/6 mice, and 2) generated polyclonal m-calpain antibodies for comparison of relative protein levels by Western blot. We found our sequence for mouse m-calpain to be almost identical to another recently published mouse sequence, and the wild-type and WLD(S) sequences to be identical. Our fusion protein and peptide polyclonal antibodies were specific for the 80 kD subunit and recognized appropriate protein bands from pure m-calpain, fusion protein, and in tissue. There was no apparent difference in m-calpain expression in nerve or spinal cord in noninjured adult animals. These data suggest that a defect in m-calpain 80 kD subunit does not likely underlie the WLD(S) phenotype, but raise questions about other subunits of calpain and possibly other proteases.
Glass
J D
JD
Department of Neurology, Emory University, Atlanta, Georgia 30322, USA. jglas03@emory.edu
Nash
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Dry
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Culver
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Levey
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Wesselingh
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D000595:Amino Acid Sequence; D000818:Animals; D001369:Axons / Q000502:physiology*; D015153:Blotting, Western; D002154:Calpain / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D007113:Immunity, Innate / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D008817:Mice, Mutant Strains / Q000235:genetics*; D008969:Molecular Sequence Data; D009410:Nerve Degeneration / Q000235:genetics*; D010446:Peptide Fragments / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D012016:Reference Values; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution
D010446:Peptide Fragments; D012333:RNA, Messenger; D002154:Calpain