title: Control of mitochondrial outer membrane permeabilization and Bcl-xL levels by thioredoxin 2 in DT40 cells.
abstract: Mitochondria play a central role in the initiation of apoptosis, which is regulated by various factors such as ATP synthesis, reactive oxygen species, redox status, and outer membrane permeabilization. Disruption of chicken thioredoxin 2 (Trx2), a mitochondrial redox-regulating protein, results in apoptosis in DT40 cells. To investigate the mechanism of this apoptosis, we prepared transfectants expressing control (DT40-TRX2-/-), human thioredoxin 2 (TRX2) (DT40-hTRX2), or redox-inactive TRX2 (DT40-hTRX2CS) in conditional Trx2-deficient DT40 cells containing a tetracycline-repressible Trx2 gene. Production of ATP was not significantly changed by down-regulation of Trx2 expression. The generation of reactive oxygen species was enhanced by the down-regulation of Trx2 expression in DT40-TRX2-/-. Unexpectedly, the change was blocked in both DT40-hTRX2 and DT40-hTRX2CS cells. The down-regulation of Trx2 expression caused the release of cytochrome c and apoptosis-inducing factor on day 3, and apoptosis on day 5. These changes were also suppressed in both DT40-hTRX2 and DT40-hTRX2CS cells, suggesting that TRX2 regulates mitochondrial outer membrane permeabilization and apoptosis by redox-active site cysteine-independent mechanisms. The down-regulation of Trx2 expression caused a decrease in the protein level of Bcl-xL on day 3, whereas the protein level of Bcl-2 did not change until day 4, and the mRNA level of Bcl-xL was unchanged. The decrease in Bcl-xL was not blocked by a caspase 3 inhibitor but blocked in both DT40-hTRX2 and DT40-hTRX2CS. These findings indicate a link between the redox active site cysteine-independent action of TRX2 and the level of Bcl-xL in the regulation of apoptosis.
authors.affiliation: Department of Biological Responses, Institute for Virus Research, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo, Kyoto 606-8507.
authors.forename: Dongmei
authors.initials: D
authors.lastname: Wang
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Masutani
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Oka
authors.forename: Toru
authors.initials: T
authors.lastname: Tanaka
authors.forename: Yuko
authors.initials: Y
authors.lastname: Yamaguchi-Iwai
authors.forename: Hajime
authors.initials: H
authors.lastname: Nakamura
authors.forename: Junji
authors.initials: J
authors.lastname: Yodoi
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D001665:Binding Sites; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D016923:Cell Death; D002460:Cell Line; D002469:Cell Separation; D002645:Chickens; D003600:Cytosol / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D015536:Down-Regulation; D005434:Flow Cytometry; D017353:Gene Deletion; D015723:Gene Library; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism*; D008564:Membrane Potentials; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism*; D010084:Oxidation-Reduction; D010920:Placenta / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017382:Reactive Oxygen Species; D013879:Thioredoxins / Q000737:chemistry* / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D051020:bcl-X Protein / Q000378:metabolism*
chemical_list: D018076:DNA, Complementary; D008565:Membrane Proteins; D017382:Reactive Oxygen Species; D051020:bcl-X Protein; D013879:Thioredoxins; D000255:Adenosine Triphosphate; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases