snoTARGET shows that human orphan snoRNA targets locate close to alternative splice junctions

Abstract

Among thousands of non-protein-coding RNAs which have been found in humans, a significant group represents snoRNA molecules that guide other types of RNAs to specific chemical modifications, cleavages, or proper folding. Yet, hundreds of mammalian snoRNAs have unknown function and are referred to as "orphan" molecules. In 2006, for the first time, it was shown that a particular orphan snoRNA (HBII-52) plays an important role in the regulation of alternative splicing of the serotonin receptor gene in humans and other mammals. In order to facilitate the investigation of possible involvement of snoRNAs in the regulation of pre-mRNA processing, we developed a new computational web resource, snoTARGET, which searches for possible guiding sites for snoRNAs among the entire set of human and rodent exonic and intronic sequences. Application of snoTARGET for finding possible guiding sites for a number of human and rodent orphan C/D-box snoRNAs showed that another subgroup of these molecules (HBII-85) have statistically elevated guiding preferences toward exons compared to introns. Moreover, these energetically favorable putative targets of HBII-85 snoRNAs are non-randomly associated with genes producing alternatively spliced mRNA isoforms. The snoTARGET resource is freely available at: (http://hsc.utoledo.edu/depts/bioinfo/snotarget.html).

Fig. 1.

Example of the snoTARGET output for a single HBII-85-12 ASE target. The second line shows the position of the 5′-end of the target inside the gene; the consecutive number of exon or intron containing the target; and the distance from the 5′-end of the target to the nearest splicing junction upstream and downstream of the target, respectively. In case the target is within the first/last exon, the distance would be to the end of the gene. The third line shows the number of G–U pairs between the ASE and the target. The fourth line shows the calculated minimum free energy between the ASE and the target (kcal/mol). The fifth line presents the target sequence itself (in the middle) and also the 30 nucleotides adjacent to it from the 5′-end and from the 3′-end, respectively. Exons are shown in uppercase while the introns in lowercase. In this example, intron #24 ends in the middle of the 5′-adjacent sequence. Finally, at the bottom, the program prints the exon/intron information of the gene containing the target, which is the entire informational line of the gene from the Exon–Intron Database (Saxonov et al., 2000).

Fig. 2.

A. Location of the ASE target relative to the alternative splice junctions in the DRF1 gene. Homo sapiens DBF4 yeast homolog B gene (GenBank mRNA identifier NM_025104). The target is present in the 3′-terminal alternative exon of isoform #3 (mRNAs NM_025104 and AK023149); while it is within alternative introns of isoforms #1 (AF448801 and BC016158) and isoform #2 (BC033660). Exons are shown as boxes, whose colored regions represent coding segments and the white region in isoform #3 represents a 3′-untranslated region. Alternative exons are shown as green boxes. The location of the target is shown by a small, red box. Exons are numbered according to isoform 16188A in the Exon–Intron Database (Saxonov et al., 2000). B. Location of the ASE target relative to the alternative splice junctions in the GTPBP3. The GTP binding protein 3 gene (UniGene identifier Hs.334885). Exon #5 (73 nt) and Exon #6 (144 nt) are shown in uppercase and are blue; retained intron #5 is shown in lowercase and is black. This intron is present in isoform IV (NM_133644) and is excluded from other isoforms (e.g. NM_032620). The target site within Exon 5 is marked in red. C. Location of the ASE target relative to the alternative splice junctions in the LRP8 gene. The low-density lipoprotein receptor-related protein 8 precursor, apolipoprotein E receptor 2 (UniGene identifier Hs.576154). The target is present inside the 854-nt-long intron 12 of the LRP8 gene. On the diagram, the constitutive exons are shown as shadowed grey boxes and the alternatively skipped exon as a green box. The location of the target is shown by a small, red box. Introns are numbered according to isoform 717A in the Exon–Intron Database (Saxonov et al., 2000). D. Scheme of the 25-nt-long ASE-1 of HBII-85-13, -14, and -16-19 snoRNAs and its target within intron 4 of the matrix metalloproteinase 16 gene (MMP16). Asterisks show U–G base-pairing. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)