Genomewide characterization of non-polyadenylated RNAs

We have used deep sequencing to explore the repertoire of both poly(A)+ and poly(A)- RNAs from HeLa cells and H9 human embryonic stem cells (hESCs). Using stringent criteria, we found that while the majority of transcripts are poly(A)+, a significant portion of transcripts are either poly(A)- or bimorphic, being found in both the poly(A)+ and poly(A)- populations. Further analyses revealed that many mRNAs may not contain classical long poly(A) tails and such messages are overrepresented in specific functional categories. In addition, we surprisingly found that a few excised introns accumulate in cells and thus constitute a new class of non-polyadenylated long non-coding RNAs. Finally, we have identified a specific subset of poly(A)- histone mRNAs, including two histone H1 variants, that are expressed in undifferentiated hESCs and are rapidly diminished upon differentiation; further, these same histone genes are induced upon reprogramming of fibroblasts to induced pluripotent stem cells.We offer a rich source of data that allows a deeper exploration of the poly(A)- landscape of the eukaryotic transcriptome. The approach we present here also applies to the analysis of the poly(A)- transcriptomes of other organisms.Nascent pre-mRNA transcripts undergo multiple co-transcriptional/post-transcriptional processing and modification events during their maturation. A poly(A) tail is added post-transcriptionally to the 3' end of almost all eukaryotic mRNAs and plays an important role in mRNA stability, nucleocytoplasmic export, and translation [1]. 3' end formation involves binding of the cleavage/polyadenylation machinery to the AAUAAA hexamer (or some variants), often together with a downstream G/U rich sequence, followed by endonucleolytic cleavage of the pre-mRNA and the addition of a 3' non-templated poly(A) tail of up to 200 to 250 adenosines in mammalian cells [2]. As most known mRNAs are polyadenylated at their 3' ends, transcriptome analysis using deep sequencing (mRN