Calpain-mediated truncation of GSK-3 in post-mortem brain samples

• Published in: Journal of neuroscience research Vol. 87; no. 5; pp. 1156 - 1161
• Main Authors: Goñi-Oliver, Paloma, Avila, Jesús, Hernández, Félix
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01.04.2009
• Subjects: Aged; Analysis of Variance; Animals; Blotting, Western; Brain - metabolism; calpain; Calpain - antagonists & inhibitors; Calpain - metabolism; Cysteine Proteinase Inhibitors - pharmacology; Dipeptides - pharmacology; Electrophoresis, Polyacrylamide Gel; Glycogen Synthase Kinase 3 - metabolism; GSK-3; Humans; Male; Mice; Mice, Inbred C57BL; Middle Aged; post-mortem; Postmortem Changes
• ISSN: 0360-4012; 1097-4547
• DOI: 10.1002/jnr.21932

GSK‐3 activity can be regulated by phosphorylation and through interaction with GSK‐3‐binding proteins. In addition, we have recently demonstrated that calpain activation produces a truncation of GSK‐3 that removes the N‐terminal inhibitory domain (Goñi‐Oliver et al. [2007] J. Biol. Chem. 282:22406). Given that calpain is involved in post‐mortem proteolysis in brain samples, the objective of this investigation was to test whether GSK‐3 is truncated in post‐mortem samples. To achieve this objective, we first investigated the degradation of GSK‐3 during different post‐mortem intervals in mouse brains and found that the conversion of GSK‐3 to proteolytic fragments of 40 and 30 kDa takes place in a way similar that of to p35‐CDK‐5 subunit and spectrin, two well‐known calpain substrates. In addition, we demonstrated that this truncation is mediated by calpain, insofar as pretreatment with MDL 28170, a permeable blood–brain barrier calpain inhibitor, partially inhibited that degradation. When human brain extracts were exposed to calcium, GSK‐3 was truncated, generating two fragments of approximately 40 and 30 kDa, a proteolytic process that was inhibited by calpeptin, a specific calpain inhibitor. Thus, this is the first report of calcium‐dependent truncation of human GSK‐3. These data demonstrate that control samples with similar post‐mortem delay are essential to interpret correctly the changes observed in GSK‐3 levels in human post‐mortem brain, especially when studying human neurodegenerative diseases. © 2008 Wiley‐Liss, Inc.