Calcium-activated neutral proteinase (calpain) activity in C6 cell line: Compartmentation of μ and m calpain

✍ Scribed by Dr. N. L. Banik; A. K. Chakrabarti; G. W. Konat; G. Gantt-Wilford; E. L. Hogan

• Publisher: John Wiley and Sons
• Year: 1992
• Tongue: English
• Weight: 669 KB
• Volume: 31
• Category: Article
• ISSN: 0360-4012
• DOI: 10.1002/jnr.490310414

✦ Synopsis

Calcium-activated neutral proteinase (calpain) activity was determined, including in cytosol and membrane fractions, in rat glioma C6 cell line. The p and m forms of calpain were separated by DEAE and phenylsepharose column chromatography and with removal of the endogenous inhibitor calpastatin. C6 cells contained more mcalpain than the p isoform. More than 70% of mcalpain activity was membraneassociated and 20 % was cytosolic. Isolated plasma membrane also contained 69% of the mcalpain activity. In contrast, approximately 80% of pcalpain activity was cytosolic and 16% was membranous. Halfmaximal activity for p and mcalpain was obtained at 1 p M and 0.2 mM CaCl,, respectively. Trypsin dissociation of cells reduced activity. Triton X-100 stimulated mcalpain activity of the whole homogenate and the membrane pellet but not of the cytosol. Activity of the myelin marker enzyme adenosine 2'3'-cyclic nucleotide 3'-phosphohydrolase (CNPase), was also found in C6 cells. The identification of calpain and CNPase in C6 cells is in keeping with an interpreta; tion that C6 differentiation resembles, at least in part, that of the myelin-forming oligodendroglial cells.