Putative conserved fern targets of <i>P</i>. <i>minima</i> miRNAs.

<p>Putative conserved fern targets of <i>P</i>. <i>minima</i> miRNAs.</p

Phylogenetic position and habit of <i>Pleopeltis minima</i>.

<p><b>(A)</b> Simplified summary of phylogenetic relationships among major plant lineages relevant for the present study [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177573#pone.0177573.ref003" target="_blank">3</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177573#pone.0177573.ref017" target="_blank">17</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177573#pone.0177573.ref024" target="_blank">24</a>]. Note that some molecular phylogenies recover a clade composed of liverworts and mosses as sister to vascular plants [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177573#pone.0177573.ref025" target="_blank">25</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177573#pone.0177573.ref026" target="_blank">26</a>]. The fern (“monilophytes”) clade is the sister group to seed plants, gymnosperms and angiosperms. Within ferns, <i>P</i>. <i>minima</i> belongs to the order Polypodiales and is more closely related to <i>Ceratopteris thalictroides</i>, while <i>Marsilea quadrifolia</i> belongs to the order Salviniales and <i>Lygodium japonicum</i> to the order Schizaeales. <b>(B)</b> Habit of <i>P</i>. <i>minima</i> growing as epiphyte upon a tree trunk. <b>(C)</b> Abaxial side of leaf surface showing form and organization. <b>(D)</b> Portion of fertile leaf showing position of sori and scales. Scale bars: 5 mm.</p

Trans-acting siRNA 3 targeting <i>ARF</i> transcripts in ferns.

<p><b>(A)</b> Schematic representations of two <i>L</i>. <i>japonicum TAS3</i> RNAs (Locus 20755 and 39179). Predicted sites targeted by pmi-miR390 are indicated in red. The region predicted to generate a tasiRNA targeting a mRNA encoding a ARF transcription factor is indicated in blue. For comparison, a <i>TAS3</i> transcript from <i>A</i>. <i>thaliana</i> (NR_143941.1) is also shown. <b>(B)</b> Schematic representation of a <i>ARF</i> mRNA (Locus 9356) from <i>L</i>. <i>japonicum</i> targeted by tasiRNA derived from <i>TAS3</i> transcripts (blue stripe). The <i>A</i>. <i>thaliana ARF3</i> mRNA (At2g33860) is shown for comparison. Conserved protein domains encoded by the <i>ARF</i> mRNAs are indicated.</p

Size distribution of <i>P</i>. <i>minima</i> sRNA sequences.

<p>The graph displays the raw number of sRNA reads in the range of 18 to 25 nt. The percentage of reads in each size category in the range is indicated. A large peak at 21 nt and a smaller peak at 24 nt are noticeable.</p

Conserved <i>P</i>. <i>minima</i> miRNAs.

<p>Conserved <i>P</i>. <i>minima</i> miRNAs.</p

Emergence of miRNAs during early land plant evolution.

<p>The schematic cladogram shows the origin of ancient miRNA families during early land plant evolution. Only miRNA families that originated in early land plant evolution are shown, namely land plants (Embryophyta, node 1), mosses and vascular plants (node 2), vascular plants (Tracheophyta, node 3) and ferns and seed plants (Euphyllophyta, node 4). Note that, according to recent molecular phylogenies [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177573#pone.0177573.ref025" target="_blank">25</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177573#pone.0177573.ref026" target="_blank">26</a>], liverworts and mosses might belong together in a clade that is sister to vascular plants, in which case nodes 1 and 2 would be the same. Also note that five miRNA families seem to be restricted to euphyllophytes and that only miR1083 seems to have originated in the branch leading to vascular plants.</p

Conserved targets of fern miRNAs.

<p>Schematic representations of <i>L</i>. <i>japonicum</i> mRNAs predicted to be targeted by <i>P</i>. <i>minima</i> miRNAs. For comparison, homologous mRNAs and miRNAs from other land plants (<i>A</i>. <i>thaliana</i> or <i>M</i>. <i>polymorpha</i>) are also shown. Coding regions are shown as rectangles and UTR regions as thick black lines. Protein domains within coding regions are coloured. Regions predicted to be targeted by pmi-miRNAs are indicated by red stripes. Note that the positions of miRNA-targeted regions in mRNAs are conserved between <i>L</i>. <i>japonicum</i> and other plants. For pmi-miR408 the sequence for the homologous <i>PPO</i> mRNA from <i>P</i>. <i>endiviifolia</i> is not available and is not shown.</p