Actinomycin D upregulates proapoptotic protein Puma and downregulates Bcl-2 mRNA in normal peripheral blood lymphocytes

作者：Kalousek Ivan^*; Brodska Barbora; Otevrelova Petra; Roselova Pavla

来源：Anti-Cancer Drugs, 2007, 18(7): 763-772.

DOI：10.1097/CAD.0b013e3280adc905

• actinomycin D
• apoptosis
• Bcl-2
• peripheral blood lymphocyte
• p53
• Puma
• ribosomal synthesis

We have examined the ability of actinomycin D to induce apoptosis in human peripheral blood lymphocytes. Run-On assays were performed to specify the primary molecular damage, reverse transcription-PCR, Western blots and flow cytometry studies were performed to ascertain which proteins of the apoptosis machinery were affected to cause actinomycin D-induced cell death. Expression of 23 apoptosis-related genes was investigated. The down-regulation of ribosomal RNA synthesis caused by actinomycin D induced a mitochondria-dependent apoptosis. Although the expression of the majority of examined genes remained indifferent against actinomycin D activity, the cellular level of p53 protein increased, subsequently upregulating both Puma mRNA and protein. Puma-mediated mitochondrial apoptosis was accompanied by nucleolin cleavage and Bcl-2 mRNA destabilization. The stability of the cellular level of Bcl-2 protein independent of a mRNA decrease suggests that protection of Bcl-2 protein against proteasomal degradation can moderate the apoptotic process. In peripheral blood lymphocytes cultured in vitro, the apoptosis induced by a low concentration of actinomycin D (10 nmol/l) is dependent on p53 and Puma activation. This apoptotic pathway is demonstrated in peripheral blood lymphocytes for the first time. A different apoptotic pathway induced in peripheral blood lymphocytes using this drug has, however, been previously revealed by other authors. The combination of cell specificity and dose-dependent effects can likely play a decisive role in apoptosis observed in peripheral blood lymphocytes after genotoxic drug application.

• 出版日期2007-8