Tumor promoter benzoyl peroxide induces sulfhydryl oxidation in protein kinase C: Its reversibility is related to the cellular resistance to peroxide-induced cytotoxicity

Citation
R. Gopalakrishna et al., Tumor promoter benzoyl peroxide induces sulfhydryl oxidation in protein kinase C: Its reversibility is related to the cellular resistance to peroxide-induced cytotoxicity, ARCH BIOCH, 363(2), 1999, pp. 246-258
Citations number
74
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemistry & Biophysics
Journal title
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
ISSN journal
0003-9861 → ACNP
Volume
363
Issue
2
Year of publication
1999
Pages
246 - 258
Database
ISI
SICI code
0003-9861(19990315)363:2<246:TPBPIS>2.0.ZU;2-W
Abstract
Since tumor promoter benzoyl peroxide (BPO) mimics phorbol esters in some a spects, its effects on protein kinase C (PKC) were previously studied, Howe ver, in those studies due to the presence of thiol agents in the PHC prepar ations, the sensitive reaction of BPO with redox-active cysteine residues i n PKC was not observed. In this study, by excluding thiol agents present in the purified PKC preparation, low concentrations of BPO modified PKC, resu lting in the loss of both kinase activity and phorbol ester binding (IC50 = 0.2 to 0.5 mu M). This modification, which was not dependent on transition metals, was totally blocked by a variety of thiol agents including OSH, wh ich directly reacted with BPO, Substoichiometric amounts of BPO (0.4 mol/mo l of PKC) oxidized two sulfhydryls in PKC and inactivated the enzyme which was readily reversed by dithiothreitol, The regulatory domain having zinc t hiolate structures supporting the membrane-inserting region provided the sp ecificity for PKC reaction with BPO, which partitioned into the membrane. U nlike H2O2, BPO did not induce the generation of the Ca2+/lipid-independent activated form of PKC, Other redox-sensitive enzymes such as protein kinas e A, phosphorylase kinase, and protein phosphatase 2A required nearly 25- t o 100-fold higher concentrations of BPO for inactivation. EPO also inactiva ted PKC in a variety of cell types. In the JB6 (30 P-) nonpromotable cell L ine and other normal cell lines, where BPO was more cytotoxic, it readily i nactivated PKC due to a slow reversibility of this inactivation by the cell . However, in the JB6 (41 P+) promotable cell line, C3H10T1/2 and B16 melan oma cells, where BPO was less cytotoxic, it did not readily inactivate PKC due to a rapid reversibility of this inactivation by an endogenous mechanis m. Nevertheless, BPO inactivated PKC at an equal rate in the homogenates pr epared from all these cell types, Inclusion of NADPH reversed this inactiva tion in the homogenates to a different extent, presumably due to a differen ce in distribution of a protein disulfide reductase, which reverses this ox idative modification. BPO-induced modification of PKC occurred independent of the cellular status of GSH, However, externally added GSH and cell-imper meable thiol agents prevented the BPO-induced modification of PHC, Since BP O readily partitions into membranes, its reaction with redox-cycling thiols of membrane proteins such as PKC may trigger epigenetic events to prevent cytotoxicity, but favor tumor promotion. (C) 1999 Academic Press.