RNA-seq analysis of normal and NMD-inhibited human erythroblasts

This experiment was designed to study a component of the erythroblast alternative splicing program that is normally masked by nonsense-mediated decay (NMD). CD34+ cells from cord blood were cultured under conditions that promote erythroblast proliferation and differentiation. RNA was isolated from proerthryoblasts / immature erythroblasts at day 9 of culture, and mature erythroblasts at day 16 of culture, and the transcriptomes of normal cells were compared with those in which NMD was inhibited by treatment with cycloheximide plus emetine. Many novel splice junctions identified in NMD-inhibited cells were located in retained introns. Experimental studies support the model that cryptic intron-internal splice sites can promote intron retention by serving as decoys that engage normal splice sites at the ends of the introns and prevent splicing at these sites.