Biochemical and biophysical research communications
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Biochem. Biophys. Res. Commun. · Feb 2000
Pyrrolidine dithiocarbamate induces apoptosis by a cytochrome c-dependent mechanism.
Pyrrolidine dithiocarbamate (PDTC) is a synthetic antioxidant molecule, which has been recently proposed as an antitumoral agent on the basis of its capability of inducing apoptosis. We investigated the effect of PDTC on the proliferation and survival of the promyelocitic cell line HL-60. Concentration as low as 10 microM of PDTC induces a significant reduction of the growth rate and the contemporaneous activation of the apoptotic process. ⋯ PDTC-dependent apoptosis was associated with an early release of cytochrome c from mitochondria, while the involvement of pathways due to cell death receptors engagement was ruled out by detailed time-course analyses of caspases 3 and 8 activation. Moreover, no up-regulation of p21(CIP1) level, a pivotal cyclin-dependent kinase inhibitor, occurred at PDTC concentration able to induce apoptosis. Finally, in vitro incubation of purified mitochondria with PDTC demonstrated that the molecule is directly able to induce cytochrome c release from the intermembrane space, thus confirming that mitochondria are a primary cellular target of the molecule.
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Biochem. Biophys. Res. Commun. · Feb 2000
Rhythmic expression of BMAL1 mRNA is altered in Clock mutant mice: differential regulation in the suprachiasmatic nucleus and peripheral tissues.
BMAL1 is a putative clock gene which encodes a basic helix-loop-helix (bHLH)-PAS transcription factor. To examine whether the CLOCK protein is required for the circadian expression of BMAL1 mRNA, in situ hybridization and Northern blot analysis were performed in the suprachiasmatic nucleus (SCN) and peripheral tissues of homozygous Clock mutant mice. In the SCN of Clock mutants, BMAL1 mRNA did not oscillate significantly but apparently expressed with low levels, while in wild-type mice the mRNA was robustly oscillated in a circadian manner. ⋯ Furthermore, expressions of BMAL1 mRNA in the peripheral of Clock mutant mice were close to the peak level in wild-type mice, whereas mPer2 mRNA levels were severely blunted at trough values. Daily expression of albumin site D-binding protein (DBP), a clock controlled output gene (CCG), was also abolished at trough values by the Clock mutation in all tissues examined. These observations suggest that the circadian expression of BMAL1 mRNA is affected by the CLOCK-induced transcriptional feedback loop in the SCN and peripheral tissues in a different way and that the regulation mechanism appeared to be different from those in mPer2 and DBP expressions in vivo.