Dz13, a DNAzyme targeting c-jun, induces off-target cytotoxicity in endothelial cells with features of nonapoptotic programmed cell death.
We have previously shown that Dz13, a catalytic DNA molecule (DNAzyme) designed against c-jun, is cytotoxic to nonquiescent cells by a mechanism independent of c-jun mRNA cleavage. In this report, we evaluated programmed cell death (PCD) pathways in order to gain further insight into the mechanism of action of Dz13. Using human dermal microvascular endothelial cells (HMEC-1), we found that Dz13-mediated cell death is characterized by mitochondrial depolarization, caspase-8 activation, lysosomal increase, and autophagosome formation. Classical DNA laddering and translocation of mitochondrial proteins were not observed. An array of inhibitors, including those targeting caspases, failed to abrogate cytotoxicity and mitochondrial depolarization. Cytotoxicity did not proceed from endoplasmic reticulum (ER) stress. The possible involvement of PARP-1 in Dz13-mediated cytotoxicity was indicated by its differential release as gauged by protein extraction data and its apparent binding to Dz13, as evidenced by protein pull-down experiments. This study on Dz13-mediated cytotoxicity presents a detailed investigation into the interplay of cell death effectors involved in apoptosis, autophagy, and necrosis, and demonstrates a novel form of oligonucleotide-mediated cytotoxicity with features of PCD.
Gozar
Mary Margaret
MM
Johnson and Johnson Research Pty Limited, Eveleigh, New South Wales, 1430, Australia. mgozar@medau.jnj.com
Goodchild
Amber
A
Passioura
Toby
T
King
Andrew
A
Lai
Angela
A
Witherington
Craig
C
Rivory
Laurent
L
D017209:Apoptosis; D001343:Autophagy; D020169:Caspases / Q000378:metabolism; D016923:Cell Death; D002460:Cell Line; D021881:DNA, Catalytic / Q000378:metabolism* / Q000494:pharmacology*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D016758:Genes, jun; D006801:Humans; D008247:Lysosomes / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D009336:Necrosis; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D014162:Transfection
D021881:DNA, Catalytic; C485475:Dz13 DNAzyme; D016755:Proto-Oncogene Proteins c-jun; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D020169:Caspases