title: Tissue transglutaminase is an in situ substrate of calpain: regulation of activity.
abstract: Tissue transglutaminase (tTG) is a calcium-dependent enzyme that catalyzes the transamidation of specific polypeptide-bound glutamine residues, a reaction that is inhibited by GTP. There is also preliminary evidence that, in situ, calpain and GTP may regulate tTG indirectly by modulating its turnover by the calcium-activated protease calpain. In the present study, the in vitro and in situ proteolysis of tTG by calpain, and modulation of this process by GTP, was examined. tTG is an excellent substrate for calpain and is rapidly degraded. Previously it has been demonstrated that GTP binding protects tTG from degradation by trypsin. In a similar manner, guanosine-5'-O-(3-thiotriphosphate) protects tTG against proteolysis by calpain. Treatment of SH-SY5Y cells with 1 nM maitotoxin, which increases intracellular calcium levels, resulted in a significant increase in in situ TG activity, with only a slight decrease in tTG protein levels. In contrast, when GTP levels were depleted by pretreating the cells with tiazofurin, maitotoxin treatment resulted in an approximately 50% decrease in tTG protein levels, and a significant decrease in TG activity, compared with maitotoxin treatment alone. Addition of calpain inhibitors inhibited the degradation of tTG in response to the combined treatment of maitotoxin and tiazofurin and resulted in a significant increase in in situ TG activity. These studies indicate that tTG is an endogenous substrate of calpain and that GTP selectively inhibits the degradation of tTG by calpain.
authors.affiliation: Department of Psychiatry and Behavioral Neurobiology, University of Alabama at Birmingham, 35294-0017, USA.
authors.forename: J
authors.initials: J
authors.lastname: Zhang
authors.forename: R P
authors.initials: RP
authors.lastname: Guttmann
authors.forename: G V
authors.initials: GV
authors.lastname: Johnson
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000494:pharmacology; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003978:Diazomethane / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004792:Enzyme Precursors / Q000494:pharmacology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006801:Humans; D008387:Marine Toxins / Q000494:pharmacology; D009447:Neuroblastoma; D009842:Oligopeptides / Q000494:pharmacology; D035964:Oxocins; D012254:Ribavirin / Q000031:analogs & derivatives / Q000494:pharmacology; D013379:Substrate Specificity; D011503:Transglutaminases / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000201:enzymology; D016875:tau Proteins / Q000378:metabolism; D000095702:Polyether Toxins
chemical_list: D000970:Antineoplastic Agents; D002118:Calcium; D002154:Calpain; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D015853:Cysteine Proteinase Inhibitors; D003978:Diazomethane; D004791:Enzyme Inhibitors; D004792:Enzyme Precursors; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D008387:Marine Toxins; D009842:Oligopeptides; D035964:Oxocins; D012254:Ribavirin; D011503:Transglutaminases; D016875:tau Proteins; C493956:CDKN1A protein, human; C090218:benzyloxycarbonylleucyl-leucyl-tyrosine diazomethyl ketone; C020394:maitotoxin; D000095702:Polyether Toxins; C098484:mu-calpain proenzyme; C033706:tiazofurin