Changes in polysome loading as a consequence of RHA downregulation [Agilent]

RNA helicase A (RHA) binds its target transcripts at the post-transcriptional control element (PCE) located in the 5’ untranslated region (UTR). This interaction represents an “RNA switch” that regulates protein synthesis. Down regulation of RHA by siRNAs was used to identify transcripts with RHA-dependent translation. Reduced accumulation of RNA in polysomes was monitored with microarrays. Cytoplasmic lysates of cells treated with RHA targeted or non-silencing control siRNAs were separated by sucrose density gradient centrifugation. Ribosomal RNA profiles were generated, containing heavy polysomes were collected, and RNA was extracted.