Estimates of the numbers of all and orthologous lincRNAs with varying expression thresholds^a.

a<p>Indel threshold: 95%, ORF threshold: 120 nt (see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#s4" target="_blank">Methods</a>). Expression thresholds were applied to lincRNA genes (Lh, Lm, and Kb) and putative orthologous lincRNA genes (Kh and Km).</p><p>% stands for conservation percentage as in <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#pcbi-1002917-t001" target="_blank">Table 1</a>.</p

RPKM-based estimates of the numbers of all and orthologous lincRNAs.

<p>Two expression and four indel thresholds were applied to putative orthologous lincRNA genes (Kh and Km).</p

The fractions of the human and mouse genomes allotted to protein-coding and lincRNA-coding sequences^a.

a<p>The data on protein-coding genes and the total size of the euchromatic genomes are from <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#pcbi.1002917-Church1" target="_blank">[31]</a>, <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#pcbi.1002917-International1" target="_blank">[63]</a>.</p>b<p>The total length of the human lincRNome is likely to be an underestimate caused by the use of RNAseq data to calculate the lengths of lincRNAs in the human validated set <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#pcbi.1002917-Cabili1" target="_blank">[27]</a>.</p

The Vast, Conserved Mammalian lincRNome

<div><p>We compare the sets of experimentally validated long intergenic non-coding (linc)RNAs from human and mouse and apply a maximum likelihood approach to estimate the total number of lincRNA genes as well as the size of the conserved part of the lincRNome. Under the assumption that the sets of experimentally validated lincRNAs are random samples of the lincRNomes of the corresponding species, we estimate the total lincRNome size at approximately 40,000 to 50,000 species, at least twice the number of protein-coding genes. We further estimate that the fraction of the human and mouse euchromatic genomes encoding lincRNAs is more than twofold greater than the fraction of protein-coding sequences. Although the sequences of most lincRNAs are much less strongly conserved than protein sequences, the extent of orthology between the lincRNomes is unexpectedly high, with 60 to 70% of the lincRNA genes shared between human and mouse. The orthologous mammalian lincRNAs can be predicted to perform equivalent functions; accordingly, it appears likely that thousands of evolutionarily conserved functional roles of lincRNAs remain to be characterized.</p> </div

The human and mouse lincRNomes.

<p>The figure shows the estimated numbers of lincRNA genes in human and mouse, and the estimate for the size of the set of orthologous lincRNAs. The circles show the estimated sizes of the human and mouse lincRNomes (<i>Nh</i> and <i>Nm</i>, respectively), and the overlap shows the estimated number of orthologous lincRNAs (<i>Nb</i>). For each of these values, the analytically determined 95% confidence intervals are indicated; bootstrap analysis yielded more narrow confidence intervals (see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#s4" target="_blank">Methods</a> for details). The small, filled circles (blue and green for human and mouse, respectively) show the validated sets of lincRNAs (<i>Lh</i> and <i>Lm</i>, respectively), and the overlap area between these circles shows orthologous expressed, validated lincRNAs (<i>Kb</i>).</p

Computational pipeline to characterize the lincRNome.

<p>The subset of orthologous lincRNAs (Kb) was obtained by comparing genomic positions of mouse and human lincRNA genes (minimal overlap 100 nucleotides), with further manual inspection of the genomic alignments. This comparison yielded 196 pairs of unique orthologous pairs of human and mouse lincRNA genes (<i>Kb</i>). Of the 4662 human lincRNAs (<i>Lh</i>), corresponding alignable regions in mouse were detected for 3529. These sequences were designated putative orthologs and checked for evidence of expression using RNAseq data for mouse tissues. Of the 3369 putative lincRNAs, for which the exon models could be determined, 2872 showed expression level greater than zero (<i>Kh</i>). Similarly, the subset of mouse lincRNAs with expressed putative orthologs (<i>Km</i>) was identified by searching for evidence of expression in human tissues. Of the 4156 mouse lincRNAs (<i>Lm</i>), for 3157 corresponding alignable regions with expression level greater than zero were identified in mouse. After applying ORF (<120 nucleotides), indel and expression thresholds (see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#s4" target="_blank">Methods</a> for details), final results (<a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#pcbi-1002917-g002" target="_blank">Figure 2</a> and <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#pcbi-1002917-t001" target="_blank">Table 1</a>) were obtained using a Maximum Likelihood Model (see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#s4" target="_blank">Methods</a> for details) and <i>Lm, Lh, Km, Kh, Kb</i> as input parameters (shown by dashed arrows) to estimate the size of the human lincRNome (<i>Nh</i>), the mouse lincRNome (<i>Nm</i>) and the orthologous subset of the two lincRNomes (<i>Nb</i>). For details of the procedures see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002917#s4" target="_blank">Methods</a>.</p