Project description:We report transcripts bound by ADAR3 through RNA immunoprecipitation of U87 cells expressing 3X-FLAG tagged ADAR3 compard to control cells
Project description:We report transcripts bound by ADAR3 through RNA immunoprecipitation of U87 cells expressing 3X-FLAG tagged ADAR3 compard to control cells
Project description:Through immunoprecipitation of epitope-tagged ADAR3 protein, several ADAR3-bound RNAs have been reported till date. However, exogenous expression of proteins are usually associated with altered gene expression profiles and thus, can influence binding profiles of RNA-binding proteins (RBPs) being studied. Here, we used affinity-purified antibody to pull down ADAR3 endogenously expressed in U373 glioblastoma cells and identified transcripts significantly enriched in the ADAR3 immunoprecipitants over control. To serve as control, antisera isolated from rabbit prior to immunization with ADAR3 antigen was used to perform the IP.
Project description:We have analyzed RNA-seq data to identify A-to-I editing sites in two groups of samples: one group isolated from human U87 cell line expressing an active ADAR3 mutant while the other isolated from U87 cell line expressing the inactive counterpart of the ADAR3 mutant. We compared these two groups of samples and identified sites whose editing levels are higher in the first group than in the second group.
Project description:Although R-domain has been implicated in RNA binding in vitro, whether ADAR3 uses the arginine-rich R-domain to mediate ADAR3-RNA interactions in vivo remain to be explored. Here, an unbiased, high-throughput approach was undertaken to understand if mutating the R-domain would affect the ability of ADAR3 to either bind and/or recognize target transcripts. Although R-domain doesn’t confer target specificity to ADAR3, the R-domain mutant is associated with decreased protein stability and might reduce the ability of ADAR3 to bind target RNAs.