Summary of Y1H results for all R2R3-MYB DNA binding domain (DBD) tested.

<p>On the left a maximum likelihood tree that defines a total of 35 DBD subgroups is displayed with collapsed branches. This tree was computed based on a multiple alignment of the R2R3 domains as defined [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0141044#pone.0141044.ref003" target="_blank">3</a>]. On top a cladogram of the 16 <i>cis</i>-elements assayed is plotted. In pink and blue are highlighted group I and group II DNA motifs, respectively. Binding scores in the matrix take values from 0 to 100% when all members of a subgroup bind to a given DNA sequence with high affinity (++ in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0141044#pone.0141044.s010" target="_blank">S4 Table</a>). Yellow square: low binding scores within DNA group I. Orange and red squares highlight the preferential binding of some DBDs toward the DNA motifs from groups IIa and IIc or groups IIa and IId, respectively.</p

Binding specificities of selected R2R3-MYBs in relation with their biological roles.

<p>Heat map representation of the Y1H results observed with selected R2R3-MYBs involved in (<b>A</b>) biotic and abiotic stress responses, (<b>B</b>) cell fate determination and flavonoid biosynthesis (in TTG1-dependent complexes) and (<b>C</b>) cell wall biosynthesis (cellulose and xylan <i>vs</i> lignins). Yellow: yeast growth on selective media, blue: no yeast growth on selective media. Stars indicate the DNA sequences that form an <i>AC</i>-rich element in between two consecutive DNA motifs (*: group IIa, **: group IId). Double head arrows indicate the most discriminating DNA motifs between the R2R3-MYB groups.</p

R2R3-MYBs binding activity

<p><b>(A)</b> Heat map representation (using the EPCLUST Tool) of the Y1H results. DNA motifs are grouped accordingly to their selectivity against the different R2R3-MYB subgroups. Yellow: yeast growth on selective media (<i>i</i>.<i>e</i>. interaction between a given DNA motif and a R2R3-MYB), blue: no yeast growth on selective media (<i>i</i>.<i>e</i>. no <i>trans</i>-activation). In pink and blue are highlighted group I and group II DNA motifs, respectively. <b>(B)</b> Box plot representation of the number of DNA motifs recognised per R2R3-MYB. <b>(C)</b> Number of DNA motifs recognised per R2R3-MYB subgroup. Error bars: binding variation amongst the R2R3-MYBs within each subgroup. Numbers above each column indicate the number of R2R3-MYB of each subgroup. S: subgroup. U: ungrouped.</p

Sequence LOGOs analysis.

<p>(<b>A</b>) Sequence LOGOs of the R2 and R3 DNA-recognition α-helices of <i>Arabidopsis thaliana</i> R2R3-MYB DBD subgroups (left) and of the bound <i>cis</i>-elements assayed on the Y1H experiments, classified into two groups (right). The bottom LOGOs are the calculated <i>consensi</i> for both groups. Boxed columns within the recognition helices highlight residues that most likely contact DNA nitrogen bases based on alignments to MYB protein data bank structures. Red arrowheads indicate key amino acid residues involved in the interaction with DNA that are conserved in almost all plant and animal MYB proteins ([<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0141044#pone.0141044.ref050" target="_blank">50</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0141044#pone.0141044.ref054" target="_blank">54</a>]). Red squares highlight key amino acid residues that are associated with specific DBD subgroups and for which some experimental evidences (<i>in vitro</i> and/or <i>in vivo</i>) on their role in the interaction with DNA targets are available. <b>(B)</b><i>AC</i>-rich sequence LOGOs associated with the <i>trans</i>-activation activity of DBD subgroup 1, 2, 3, 13 and 24.</p

Hierarchical clustering analysis of the binding scores associated with the 35 DNA-binding domains toward the DNA motifs belonging to group II.

<p>Preferential binding of DBDs toward groups IIa and IIc or groups IIa and IId are highlighted by orange and red squares, respectively. Stars indicate the DNA sequences that form an <i>AC</i>-rich element in between two consecutive DNA motifs (*:group IIa, **: group IId). Arrowheads indicate R2R3-MYB DBDs that were found to be the most strongly associated with group IIb (<i>AC</i>-elements) DNA motifs in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0141044#pone.0141044.g003" target="_blank">Fig 3</a>.</p