Abstract
We have developed an original vector library that allowed us to exploit the phenomenon of RNA interference but also allowed us to avoid the confounding effects of the interferon response. In the present work, we used our library of small interfering RNA expression vectors to examine the genes involved in apoptosis that was induced by double-stranded RNA. To our surprise, screening of our library revealed two novel double-stranded RNA-induced apoptotic pathways, a JNK/SAPK-mediated mitochondrial pathway and an ERK2-related pathway, both of which appeared to be independent of the serine-threonine protein kinase-dependent caspase pathway. We also found that MST2 and protein kinase Calpha both activated the pro-apoptotic signal mediated by ERK2. The results of our screening analysis suggested the utility of large scale screenings with libraries of small interfering RNA expression vectors.
Highlights
RNA interference (RNAi),1 reported originally by Fire et al (1), is a phenomenon whereby small double-stranded RNAs induce the degradation of corresponding target mRNAs
Several groups, including our own, have circumvented these problems by developing systems for vector-based RNAi (4 –10). This approach enables the maintenance of RNAi activity for much longer periods than can be achieved with synthesized small interfering RNAs (siRNAs), and siRNA expression vectors increase the possibility of the application of RNAi as a practical approach to gene silencing
The induction of apoptosis by double-stranded RNAs (dsRNAs) is remarkable, with respect to the interferon response that is associated with RNAi and as a mechanism by which mammalian cells protect themselves against viral infection
Summary
RNA interference (RNAi),1 reported originally by Fire et al (1), is a phenomenon whereby small double-stranded RNAs (dsRNAs) induce the degradation of corresponding target mRNAs. The two types of stable cells were transfected with siRNA expression vectors targeted against Renilla luciferase (negative control) or ERK2, respectively. The involvement of Bid, BAX, VDAC, cytochrome c, caspase 9, and Apaf-1 suggests that the mitochondrial apoptotic pathway might play an important role in dsRNA-induced apoptosis.
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