A comprehensive phylogeny of auxin homeostasis genes involved in adventitious root formation in carnation stem cuttings

<div><p>Understanding the functional basis of auxin homeostasis requires knowledge about auxin biosynthesis, auxin transport and auxin catabolism genes, which is not always directly available despite the recent whole-genome sequencing of many plant species. Through sequence homology searches and phylogenetic analyses on a selection of 11 plant species with high-quality genome annotation, we identified the putative gene homologs involved in auxin biosynthesis, auxin catabolism and auxin transport pathways in carnation (<i>Dianthus caryophyllus</i> L.). To deepen our knowledge of the regulatory events underlying auxin-mediated adventitious root formation in carnation stem cuttings, we used RNA-sequencing data to confirm the expression profiles of some auxin homeostasis genes during the rooting of two carnation cultivars with different rooting behaviors. We also confirmed the presence of several auxin-related metabolites in the stem cutting tissues. Our findings offer a comprehensive overview of auxin homeostasis genes in carnation and provide a solid foundation for further experiments investigating the role of auxin homeostasis in the regulation of adventitious root formation in carnation.</p></div

Auxin biosynthesis protein families in carnation.

<p>Auxin biosynthesis protein families in carnation.</p

Auxin biosynthesis protein families in carnation.

<p>Auxin biosynthesis protein families in carnation.</p

Auxin catabolism protein families in carnation.

<p>Auxin catabolism protein families in carnation.</p

Auxin transport protein families in carnation.

<p>Auxin transport protein families in carnation.</p

Proposed pathways of IAA homeostasis in carnation stem cuttings.

<p>(A) Auxin biosynthesis. In the middle box, the IPyA pathway is shown, while the IAM pathway is shown in the left-side box. The IAOx pathway, which is restricted to indole-glucosinolate-producing plant species such as <i>Arabidopsis thaliana</i>, <i>Brassica napus</i> or <i>Sinapis alba</i>, is given in the right-side box. (B) Auxin transport. (C) Auxin catabolism. Dashed lines indicate assumed reaction steps for which the corresponding enzymes have yet to be identified. Proteins are abbreviated as follows: ABCB, ATP-binding cassette transporter subfamily B; AMI1, amidase 1; AUX1/LAX, AUX1 and LAX auxin influx carriers; CYP79B2/B3, cytochrome P450 monooxygenase 79B2/B3; DAO, dioxygenase for auxin oxidation; GH3, GRETCHEN HAGEN3 acyl amido synthase; IAMT, indole-3-acetic acid methyltransferase; NIT, nitrilase; PIN, PIN-FORMED auxin efflux facilitators; UGT (group L), UDP glucosyltransferases; TAA1, tryptophan aminotransferase of Arabidopsis 1; TAR, tryptophan aminotransferase-related; YUC, YUCCA. Metabolites are abbreviated as follows: IAA, indole-3-acetic acid; IAA-Asp, indole-3-acetyl-aspartic acid; IAA-Glc, 1-O-(indol-3-ylacetyl)-β-D-glucose; IAM, indol-3-acetamide; IAN, indole-3-acetonitrile; IAOx. indole-3-acetaldoxime; IPyA, indole-3-pyruvic acid; MeIAA, methyl-IAA ester; oxIAA, oxindole-3-IAA; Trp, tryptophan.</p

Auxin-related metabolites (ng g^-1 fresh weight) identified in carnation stem cuttings.

<p>Auxin-related metabolites (ng g^-1 fresh weight) identified in carnation stem cuttings.</p

Phylogenetic analyses of the TAA1/TAR gene family in carnation.

<p>Boxes show magnifications of the tree branches containing the (A-C) TAA1/TAR protein family members studied in this work. Full phylogenetic trees are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196663#pone.0196663.s002" target="_blank">S1 Fig</a>. The evolutionary history was inferred by using the Maximum Likelihood method based on the Le Gascuel model [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196663#pone.0196663.ref038" target="_blank">38</a>]. Trees were drawn to scale, with branch lengths representing the number of substitutions per site. These analyses were conducted in MEGA7 [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196663#pone.0196663.ref029" target="_blank">29</a>], as described in the Materials and methods section. (D) Gene structure and (E) expression profiles of TAA1/TAR genes. UTR regions and exons are represented by white and black boxes, respectively; introns are depicted as gray lines. UTR, intron and exon lengths were determined using previous RNA-seq data [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196663#pone.0196663.ref006" target="_blank">6</a>].</p

Phylogenetic analyses of the AMI1 gene family in carnation.

<p>(A) Magnification of the tree branch containing the AMI1 protein family members studied in this work. The full phylogenetic tree is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196663#pone.0196663.s004" target="_blank">S3 Fig</a>. (B) Gene structure and (C) gene expression profiles of AMI1 genes. See the legend in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196663#pone.0196663.g002" target="_blank">Fig 2</a> for details.</p

Phylogenetic analyses of the YUC gene family in carnation.

<p>Boxes show the magnification of tree branches containing the (A-D) YUC protein family members studied in this work. Full phylogenetic trees are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196663#pone.0196663.s003" target="_blank">S2 Fig</a>. (E) Gene structure of YUC genes. See legend in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196663#pone.0196663.g002" target="_blank">Fig 2</a> for details.</p