Genome-wide RNAi Screening to Identify Host Factors That Modulate Oncolytic Virus Therapy.

High-throughput genome-wide RNAi (RNA interference) screening technology has been widely used for discovering host factors that impact virus replication. Here we present the application of this technology to uncovering host targets that specifically modulate the replication of Maraba virus, an oncolytic rhabdovirus, and vaccinia virus with the goal of enhancing therapy. While the protocol has been tested for use with oncolytic Maraba virus and oncolytic vaccinia virus, this approach is applicable to other oncolytic viruses and can also be utilized for identifying host targets that modulate virus replication in mammalian cells in general.

Inhibitor of apoptosis protein cIAP2 is essential for lipopolysaccharide-induced macrophage survival.

The cellular inhibitor of apoptosis 2 (cIAP2/HIAP1) is a potent inhibitor of apoptotic death. In contrast to the other members of the IAP family, cIAP2 is transcriptionally inducible by nuclear factor-kappaB in response to multiple triggers. We demonstrate here that cIAP2-/- mice exhibit profound resistance to lipopolysaccharide (LPS)-induced sepsis, specifically because of an attenuated inflammatory response.

Spurious splicing within the XIAP 5' UTR occurs in the Rluc/Fluc but not the betagal/CAT bicistronic reporter system.

X-chromosome-linked inhibitor of apoptosis, XIAP, has been shown to contain a strong internal ribosome entry site (IRES) within its 5' untranslated region (UTR) that promotes translation of XIAP mRNA under conditions of cellular stress. This claim came under scrutiny in a recent report demonstrating that the XIAP 5' UTR undergoes splicing when inserted between the two reporter cistrons of the dual luciferase plasmid Rluc/Fluc. In this paper, we demonstrate that the splicing within the XIAP 5' UTR specifically occurs only in the context of mRNA produced from the Rluc/Fluc but not the pbetagal/CAT bicistronic reporter plasmid.

Loss of XIAP protein expression by RNAi and antisense approaches sensitizes cancer cells to functionally diverse chemotherapeutics.

Stable expression of short-hairpin RNAs (shRNAs) directed against the X-linked inhibitor of apoptosis (XIAP) resulted in the generation of three MDA-MB-231 cell lines (XIAP shRNA cells) with reductions in XIAP mRNA and protein levels > 85% relative to MDA-MB-231 cells stably transfected with the U6 RNA polymerase III promoter alone (U6 cells). This RNA interference (RNAi) approach dramatically sensitized these cells to killing by the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Importantly, loss of XIAP also sensitized the cells to killing by taxanes but had no additional effects on killing by carboplatin and doxorubicin.

Cloning and characterization of the rat homologues of the Inhibitor of Apoptosis protein 1, 2, and 3 genes.

Background: Inhibitor of Apoptosis (IAP) proteins are key intrinsic regulators of apoptosis induced by a variety of triggers. We isolated the rat Inhibitor of Apoptosis genes 1, 2 and 3 and characterized their tissue distribution and expression.

Results: Rat iap-1 encodes a protein of 67.