The transcriptomes of eukaryotic cells are incredibly complex. Individual non-coding RNAs dwarf the number of protein-coding genes, and include classes that are well understood as well as classes for which the nature, extent and functional roles are obscure1. Deep sequencing of small RNAs (<200 nucleotides) from human HeLa and HepG2 cells revealed a remarkable breadth of species. These arose both from within annotated genes and from unannotated intergenic regions. Overall, small RNAs tended to align with CAGE (cap-analysis of gene expression) tags2, which mark the 5' ends of capped, long RNA transcripts. Many small RNAs, including the previously described promoter-associated small RNAs3, appeared to possess cap structures. Members of an extensive class of both small RNAs and CAGE tags were distributed across internal exons of annotated protein coding and non-coding genes, sometimes crossing exon–exon junctions. Here we show that processing of mature mRNAs through an as yet unknown mechanism may generate complex populations of both long and short RNAs whose apparently capped 5' ends coincide. Supplying synthetic promoter-associated small RNAs corresponding to the c-MYC transcriptional start site reduced MYC messenger RNA abundance. The studies presented here expand the catalogue of cellular small RNAs and demonstrate a biological impact for at least one class of non-canonical small RNAs.

The repertoire of RNAs found in eukaryotic cells is unexpectedly complex, with virtually the entire non-repeat portions of many genomes being transcribed1. Genic regions are often populated by interleaved transcription units, which give rise to both protein-coding RNAs and long and short non-coding RNAs1. Promoter-associated short RNAs (PASRs) and termini-associated short RNAs (TASRs) are recent additions to the pantheon of short RNAs4. Although their functions are unknown, several of their characteristics support biological significance. For example, PASRs and TASRs cluster at 5' and 3' termini of annotated genes4. Overall, the presence of PASRs correlates with the expression of a given locus, but not all expressed loci generate these species. Moreover, the production of PASRs and TASRs from particular loci is a conserved feature of the human and mouse genomes4. As part of our ongoing effort to understand the full repertoire of small RNAs, their mechanisms of biogenesis and their biological impacts, we analysed the small RNAs (<200 nucleotides (nt)) of HepG2 and HeLa cell lines using next-generation sequencing5.