Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1997 Jul 15;325(Pt 2):315–319. doi: 10.1042/bj3250315

Bcl-xL overexpression attenuates glutathione depletion in FL5.12 cells following interleukin-3 withdrawal.

H K Bojes 1, K Datta 1, J Xu 1, A Chin 1, P Simonian 1, G Nuñez 1, J P Kehrer 1
PMCID: PMC1218562  PMID: 9230108

Abstract

Bcl-xL and bax are bcl-2-related genes whose protein products either inhibit or promote apoptosis. Oxidative damage, including the loss of glutathione, has been implicated in the induction of apoptosis. The ability of the Bcl proteins to affect GSH was assessed in control, bax- and bcl-xL-transfected FL5.12 cells [an interleukin (IL)-3-dependent murine prolymphocytic cell line]. Overall levels of GSH were approximately the same in control and bcl-xL transfectants during the 6 h incubation period, although levels increased in bcl-xL transfectants 24 h after replating. GSH in cells overexpressing bax was reduced by approximately 36%. There were no consistent differences between these cell lines in the activities of superoxide dismutase, catalase, glutathione peroxidase or glutathione reductase. Following IL-3 withdrawal, a condition known to cause apoptosis in these cells, a rapid loss of intracellular GSH occurred in control and bax transfectants, which preceded the onset of apoptosis. GSH depletion could not be attributed to intracellular oxidation but rather seemed to occur due to a translocation out of the cell. Cells overexpressing bcl-xL did not lose significant amounts of GSH upon withdrawal of IL-3, and no apoptosis was evident. These results suggest a possible role for GSH in the mechanism by which bcl-xL prevents cell death.

Full Text

The Full Text of this article is available as a PDF (343.2 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beaver J. P., Waring P. A decrease in intracellular glutathione concentration precedes the onset of apoptosis in murine thymocytes. Eur J Cell Biol. 1995 Sep;68(1):47–54. [PubMed] [Google Scholar]
  2. Boise L. H., González-García M., Postema C. E., Ding L., Lindsten T., Turka L. A., Mao X., Nuñez G., Thompson C. B. bcl-x, a bcl-2-related gene that functions as a dominant regulator of apoptotic cell death. Cell. 1993 Aug 27;74(4):597–608. doi: 10.1016/0092-8674(93)90508-n. [DOI] [PubMed] [Google Scholar]
  3. Bustamante J., Tovar-B A., Montero G., Boveris A. Early redox changes during rat thymocyte apoptosis. Arch Biochem Biophys. 1997 Jan 1;337(1):121–128. doi: 10.1006/abbi.1996.9754. [DOI] [PubMed] [Google Scholar]
  4. Carlberg I., Mannervik B. Glutathione reductase. Methods Enzymol. 1985;113:484–490. doi: 10.1016/s0076-6879(85)13062-4. [DOI] [PubMed] [Google Scholar]
  5. Chao D. T., Linette G. P., Boise L. H., White L. S., Thompson C. B., Korsmeyer S. J. Bcl-XL and Bcl-2 repress a common pathway of cell death. J Exp Med. 1995 Sep 1;182(3):821–828. doi: 10.1084/jem.182.3.821. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Corcoran G. B., Fix L., Jones D. P., Moslen M. T., Nicotera P., Oberhammer F. A., Buttyan R. Apoptosis: molecular control point in toxicity. Toxicol Appl Pharmacol. 1994 Oct;128(2):169–181. doi: 10.1006/taap.1994.1195. [DOI] [PubMed] [Google Scholar]
  7. Ellerby L. M., Ellerby H. M., Park S. M., Holleran A. L., Murphy A. N., Fiskum G., Kane D. J., Testa M. P., Kayalar C., Bredesen D. E. Shift of the cellular oxidation-reduction potential in neural cells expressing Bcl-2. J Neurochem. 1996 Sep;67(3):1259–1267. doi: 10.1046/j.1471-4159.1996.67031259.x. [DOI] [PubMed] [Google Scholar]
  8. Fang W., Nath K. A., Mackey M. F., Noelle R. J., Mueller D. L., Behrens T. W. CD40 inhibits B cell apoptosis by upregulating bcl-xL expression and blocking oxidant accumulation. Am J Physiol. 1997 Mar;272(3 Pt 1):C950–C956. doi: 10.1152/ajpcell.1997.272.3.C950. [DOI] [PubMed] [Google Scholar]
  9. González-García M., Pérez-Ballestero R., Ding L., Duan L., Boise L. H., Thompson C. B., Núez G. bcl-XL is the major bcl-x mRNA form expressed during murine development and its product localizes to mitochondria. Development. 1994 Oct;120(10):3033–3042. doi: 10.1242/dev.120.10.3033. [DOI] [PubMed] [Google Scholar]
  10. Harlan J. M., Levine J. D., Callahan K. S., Schwartz B. R., Harker L. A. Glutathione redox cycle protects cultured endothelial cells against lysis by extracellularly generated hydrogen peroxide. J Clin Invest. 1984 Mar;73(3):706–713. doi: 10.1172/JCI111263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hockenbery D. M., Oltvai Z. N., Yin X. M., Milliman C. L., Korsmeyer S. J. Bcl-2 functions in an antioxidant pathway to prevent apoptosis. Cell. 1993 Oct 22;75(2):241–251. doi: 10.1016/0092-8674(93)80066-n. [DOI] [PubMed] [Google Scholar]
  12. Hockenbery D., Nuñez G., Milliman C., Schreiber R. D., Korsmeyer S. J. Bcl-2 is an inner mitochondrial membrane protein that blocks programmed cell death. Nature. 1990 Nov 22;348(6299):334–336. doi: 10.1038/348334a0. [DOI] [PubMed] [Google Scholar]
  13. Hug H., Enari M., Nagata S. No requirement of reactive oxygen intermediates in Fas-mediated apoptosis. FEBS Lett. 1994 Sep 12;351(3):311–313. doi: 10.1016/0014-5793(94)00852-3. [DOI] [PubMed] [Google Scholar]
  14. Ishikawa T., Ali-Osman F. Glutathione-associated cis-diamminedichloroplatinum(II) metabolism and ATP-dependent efflux from leukemia cells. Molecular characterization of glutathione-platinum complex and its biological significance. J Biol Chem. 1993 Sep 25;268(27):20116–20125. [PubMed] [Google Scholar]
  15. Ishikawa T. The ATP-dependent glutathione S-conjugate export pump. Trends Biochem Sci. 1992 Nov;17(11):463–468. doi: 10.1016/0968-0004(92)90489-v. [DOI] [PubMed] [Google Scholar]
  16. Kane D. J., Sarafian T. A., Anton R., Hahn H., Gralla E. B., Valentine J. S., Ord T., Bredesen D. E. Bcl-2 inhibition of neural death: decreased generation of reactive oxygen species. Science. 1993 Nov 19;262(5137):1274–1277. doi: 10.1126/science.8235659. [DOI] [PubMed] [Google Scholar]
  17. Kluck R. M., Bossy-Wetzel E., Green D. R., Newmeyer D. D. The release of cytochrome c from mitochondria: a primary site for Bcl-2 regulation of apoptosis. Science. 1997 Feb 21;275(5303):1132–1136. doi: 10.1126/science.275.5303.1132. [DOI] [PubMed] [Google Scholar]
  18. Krajewski S., Tanaka S., Takayama S., Schibler M. J., Fenton W., Reed J. C. Investigation of the subcellular distribution of the bcl-2 oncoprotein: residence in the nuclear envelope, endoplasmic reticulum, and outer mitochondrial membranes. Cancer Res. 1993 Oct 1;53(19):4701–4714. [PubMed] [Google Scholar]
  19. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  20. Lee J. C., Hapel A. J., Ihle J. N. Constitutive production of a unique lymphokine (IL 3) by the WEHI-3 cell line. J Immunol. 1982 Jun;128(6):2393–2398. [PubMed] [Google Scholar]
  21. Liu H., Kehrer J. P. The reduction of glutathione disulfide produced by t-butyl hydroperoxide in respiring mitochondria. Free Radic Biol Med. 1996;20(3):433–442. doi: 10.1016/0891-5849(95)02093-4. [DOI] [PubMed] [Google Scholar]
  22. Liu X., Kim C. N., Yang J., Jemmerson R., Wang X. Induction of apoptotic program in cell-free extracts: requirement for dATP and cytochrome c. Cell. 1996 Jul 12;86(1):147–157. doi: 10.1016/s0092-8674(00)80085-9. [DOI] [PubMed] [Google Scholar]
  23. Martin S. J., Green D. R. Protease activation during apoptosis: death by a thousand cuts? Cell. 1995 Aug 11;82(3):349–352. doi: 10.1016/0092-8674(95)90422-0. [DOI] [PubMed] [Google Scholar]
  24. Minn A. J., Vélez P., Schendel S. L., Liang H., Muchmore S. W., Fesik S. W., Fill M., Thompson C. B. Bcl-x(L) forms an ion channel in synthetic lipid membranes. Nature. 1997 Jan 23;385(6614):353–357. doi: 10.1038/385353a0. [DOI] [PubMed] [Google Scholar]
  25. Muchmore S. W., Sattler M., Liang H., Meadows R. P., Harlan J. E., Yoon H. S., Nettesheim D., Chang B. S., Thompson C. B., Wong S. L. X-ray and NMR structure of human Bcl-xL, an inhibitor of programmed cell death. Nature. 1996 May 23;381(6580):335–341. doi: 10.1038/381335a0. [DOI] [PubMed] [Google Scholar]
  26. Muschel R. J., Bernhard E. J., Garza L., McKenna W. G., Koch C. J. Induction of apoptosis at different oxygen tensions: evidence that oxygen radicals do not mediate apoptotic signaling. Cancer Res. 1995 Mar 1;55(5):995–998. [PubMed] [Google Scholar]
  27. Neuschwander-Tetri B. A., Roll F. J. Glutathione measurement by high-performance liquid chromatography separation and fluorometric detection of the glutathione-orthophthalaldehyde adduct. Anal Biochem. 1989 Jun;179(2):236–241. doi: 10.1016/0003-2697(89)90121-8. [DOI] [PubMed] [Google Scholar]
  28. Oltvai Z. N., Milliman C. L., Korsmeyer S. J. Bcl-2 heterodimerizes in vivo with a conserved homolog, Bax, that accelerates programmed cell death. Cell. 1993 Aug 27;74(4):609–619. doi: 10.1016/0092-8674(93)90509-o. [DOI] [PubMed] [Google Scholar]
  29. Oude Elferink R. P., Ottenhoff R., Radominska A., Hofmann A. F., Kuipers F., Jansen P. L. Inhibition of glutathione-conjugate secretion from isolated hepatocytes by dipolar bile acids and other organic anions. Biochem J. 1991 Feb 15;274(Pt 1):281–286. doi: 10.1042/bj2740281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Payne C. M., Bernstein C., Bernstein H. Apoptosis overview emphasizing the role of oxidative stress, DNA damage and signal-transduction pathways. Leuk Lymphoma. 1995 Sep;19(1-2):43–93. doi: 10.3109/10428199509059662. [DOI] [PubMed] [Google Scholar]
  31. Sandstrom P. A., Pardi D., Tebbey P. W., Dudek R. W., Terrian D. M., Folks T. M., Buttke T. M. Lipid hydroperoxide-induced apoptosis: lack of inhibition by Bcl-2 over-expression. FEBS Lett. 1995 May 22;365(1):66–70. doi: 10.1016/0014-5793(95)00443-d. [DOI] [PubMed] [Google Scholar]
  32. Shimizu S., Eguchi Y., Kosaka H., Kamiike W., Matsuda H., Tsujimoto Y. Prevention of hypoxia-induced cell death by Bcl-2 and Bcl-xL. Nature. 1995 Apr 27;374(6525):811–813. doi: 10.1038/374811a0. [DOI] [PubMed] [Google Scholar]
  33. Simonian P. L., Grillot D. A., Merino R., Nuñez G. Bax can antagonize Bcl-XL during etoposide and cisplatin-induced cell death independently of its heterodimerization with Bcl-XL. J Biol Chem. 1996 Sep 13;271(37):22764–22772. doi: 10.1074/jbc.271.37.22764. [DOI] [PubMed] [Google Scholar]
  34. Slater A. F., Stefan C., Nobel I., van den Dobbelsteen D. J., Orrenius S. Intracellular redox changes during apoptosis. Cell Death Differ. 1996 Jan;3(1):57–62. [PubMed] [Google Scholar]
  35. Thompson C. B. Apoptosis in the pathogenesis and treatment of disease. Science. 1995 Mar 10;267(5203):1456–1462. doi: 10.1126/science.7878464. [DOI] [PubMed] [Google Scholar]
  36. Vaux D. L., Cory S., Adams J. M. Bcl-2 gene promotes haemopoietic cell survival and cooperates with c-myc to immortalize pre-B cells. Nature. 1988 Sep 29;335(6189):440–442. doi: 10.1038/335440a0. [DOI] [PubMed] [Google Scholar]
  37. Veis D. J., Sorenson C. M., Shutter J. R., Korsmeyer S. J. Bcl-2-deficient mice demonstrate fulminant lymphoid apoptosis, polycystic kidneys, and hypopigmented hair. Cell. 1993 Oct 22;75(2):229–240. doi: 10.1016/0092-8674(93)80065-m. [DOI] [PubMed] [Google Scholar]
  38. Watson R. W., Rotstein O. D., Nathens A. B., Dackiw A. P., Marshall J. C. Thiol-mediated redox regulation of neutrophil apoptosis. Surgery. 1996 Aug;120(2):150–158. doi: 10.1016/s0039-6060(96)80282-0. [DOI] [PubMed] [Google Scholar]
  39. Yang J., Liu X., Bhalla K., Kim C. N., Ibrado A. M., Cai J., Peng T. I., Jones D. P., Wang X. Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked. Science. 1997 Feb 21;275(5303):1129–1132. doi: 10.1126/science.275.5303.1129. [DOI] [PubMed] [Google Scholar]
  40. Zhong L. T., Sarafian T., Kane D. J., Charles A. C., Mah S. P., Edwards R. H., Bredesen D. E. bcl-2 inhibits death of central neural cells induced by multiple agents. Proc Natl Acad Sci U S A. 1993 May 15;90(10):4533–4537. doi: 10.1073/pnas.90.10.4533. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. van den Dobbelsteen D. J., Nobel C. S., Schlegel J., Cotgreave I. A., Orrenius S., Slater A. F. Rapid and specific efflux of reduced glutathione during apoptosis induced by anti-Fas/APO-1 antibody. J Biol Chem. 1996 Jun 28;271(26):15420–15427. doi: 10.1074/jbc.271.26.15420. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES