RNomics: an experimental approach that identifies 201 candidates for novel, small, non-messenger RNAs in mouse

Abstract

In mouse brain cDNA libraries generated from small RNA molecules we have identified a total of 201 different expressed RNA sequences potentially encoding novel small non‐messenger RNA species (snmRNAs). Based on sequence and structural motifs, 113 of these RNAs can be assigned to the C/D box or H/ACA box subclass of small nucleolar RNAs (snoRNAs), known as guide RNAs for rRNA. While 30 RNAs represent mouse homologues of previously identified human C/D or H/ACA snoRNAs, 83 correspond to entirely novel snoRNAs. Among these, for the first time, we identified four C/D box snoRNAs and four H/ACA box snoRNAs predicted to direct modifications within U2, U4 or U6 small nuclear RNAs (snRNAs). Furthermore, 25 snoRNAs from either class lacked antisense elements for rRNAs or snRNAs. Therefore, additional snoRNA targets have to be considered. Surprisingly, six C/D box snoRNAs and one H/ACA box snoRNA were expressed exclusively in brain. Of the 88 RNAs not belonging to either snoRNA subclass, at least 26 are probably derived from truncated heterogeneous nuclear RNAs (hnRNAs) or mRNAs. Short interspersed repetitive elements (SINEs) are located on five RNA sequences and may represent rare examples of transcribed SINEs. The remaining RNA species could not as yet be assigned either to any snmRNA class or to a part of a larger hnRNA/mRNA. It is likely that at least some of the latter will represent novel, unclassified snmRNAs.

Complete sequencing and characterization of 21,243 full-length human cDNAs

“…4 online). Although some of these cDNAs might be cloning artifacts, recent evidence indicates that unexpectedly large populations of non-protein-coding transcripts exist in mammalian cells 16,17 . Genomic alignments of these clusters clearly showed that splicing had occurred for 1,378 of the 5,481 clusters.…”

Pseudouridines in spliceosomal snRNAs

“…To test this idea, several labs focused their resources into a search for RNA guide sequences that matched those of spliceosomal snRNAs, and successfully found a number of candidate box H/ACA guide RNAs (Hüttenhofer et al, 2001;Jady and Kiss, 2001;Hüttenhofer et al, 2002;Zhao et al, 2002;Schattner et al, 2006). Detailed experimental analyses, using the mammalian system as well as the Xenopus oocyte microinjection system, confirmed the presence of guide activity stemming from a couple of the identified box H/ACA RNAs (Jády and Kiss, 2001;Zhao et al, 2002).…”

“…To date, the systematic research using a variety of sources, including computational predictions and experimental approaches, has led to the identification of an almostcomplete library of box H/ACA guide RNAs corresponding to nearly all sites of pseudouridylation in higher eukaryotic rRNAs and snRNAs (Hüttenhofer et al, 2001;Schattner et al, 2006). These results suggest that, in higher eukaryotes, RNAguided RNA modification is the main (if not the only) mechanism for the pseudouridylation of rRNAs and spliceosomal snRNAs.…”

Evolutionary patterns of non-coding RNAs

“…A recent survey of the slime mold Dictyostelium discoideum uncovered two novel classes of ncRNAs [12]. An experimental screen recovered hundreds of small ncRNAs from the mouse [175]. Ambros and coworkers [7] reported more than 30 tiny non-coding RNAs in a recent survey of Caenorhabditis elegans that are slightly shorter than microRNAs, are not processed from hairpin precursors, and are poorly conserved between related species.…”