Co-immunoprecipitation of RNAs with RNA helicase A (RHA)

Given that RHA regulates translation by binding a PCE located at the 5’ UTR of the target transcripts a genome-wide screen was performed to identify mRNAs that bind RHA in vivo. The results from duplicate experiments with samples obtained in two independent RNA immunoprecipitations identified 407 transcripts that co-immunoprecipitate with FLAG RHA. Keywords: gene expression array-based Since N-terminal FLAG tagged RHA specifically co-immunoprecipitates PCE-containing mRNAs COS cells were transfected with a CMV-FLAG-RHA construct. Cytoplasmic lysates were pooled and immunoprecipitated with anti-FLAG beads. RNA was extracted from the immunoprecipitate and used to probe rhesus macaque Affimetrix expression arrays. Steady state mRNA levels were monitored on a separate array probed with total RNA.