The family of DOF transcription factors in Brachypodium distachyon: phylogenetic comparison with rice and barley DOFs and expression profiling

ABSTRACT: Transcription factors (TFs) are proteins that have played a central role both in evolution and in domestication, and are major regulators of development in living organisms. Plant genome sequences reveal that approximately 7% of all genes encode putative TFs. The DOF (DNA binding with One Finger) TF family has been associated with vital processes exclusive to higher plants and to their close ancestors (algae, mosses and ferns). These are seed maturation and germination, light-mediated regulation, phytohormone and plant responses to biotic and abiotic stresses, etc. In Hordeum vulgare and Oryza sativa, 26 and 30 different Dof genes, respectively, have been annotated. Brachypodium distachyon has been the first Pooideae grass to be sequenced and, due to its genomic, morphological and physiological characteristics, has emerged as the model system for temperate cereals, such as wheat and barley. RESULTS: Through searches in the B. distachyon genome, 27 Dof genes have been identified and a phylogenetic comparison with the Oryza sativa and the Hordeum vulgare DOFs has been performed. To explore the evolutionary relationship among these DOF proteins, a combined phylogenetic tree has been constructed with the Brachypodium DOFs and those from rice and barley. This phylogenetic analysis has classified the DOF proteins into four Major Cluster of Orthologous Groups (MCOGs). Using RT-qPCR analysis the expression profiles of the annotated BdDof genes across four organs (leaves, roots, spikes and seeds) has been investigated. These results have led to a classification of the BdDof genes into two groups, according to their expression levels. The genes highly or preferentially expressed in seeds have been subjected to a more detailed expression analysis (maturation, dry stage and germination). CONCLUSIONS: Comparison of the expression profiles of the Brachypodium Dof genes with the published functions of closely related DOF sequences from the cereal species considered here, deduced from the phylogenetic analysis, indicates that although the expression profile has been conserved in many of the putative orthologs, in some cases duplication followed by subsequent divergence may have occurred (neo-functionalization).

Arabidopsis DELLA and two HD-ZIP transcription factors regulate GA signalling in the epidermis through the L1 cis-element

Gibberellins (GAs) are plant hormones that affect plant growth and regulate gene expression differentially across tissues. To study the molecular mechanisms underlying GA signaling in Arabidopsis thaliana, we focused on a GDSL lipase gene (LIP1) induced by GA and repressed by DELLA proteins. LIP1 contains an L1 box promoter sequence, conserved in the promoters of epidermis-specific genes, that is bound by ATML1, an HD-ZIP transcription factor required for epidermis specification. In this study, we demonstrate that LIP1 is specifically expressed in the epidermis and that its L1 box sequence mediates GA-induced transcription. We show that this sequence is overrepresented in the upstream regulatory regions of GA-induced and DELLA-repressed transcriptomes and that blocking GA signaling in the epidermis represses the expression of L1 box–containing genes and negatively affects seed germination. We show that DELLA proteins interact directly with ATML1 and its paralogue PDF2 and that silencing of both HD-ZIP transcription factors inhibits epidermal gene expression and delays germination. Our results indicate that, upon seed imbibition, increased GA levels reduce DELLA protein abundance and release ATML1/PDF2 to activate L1 box gene expression, thus enhancing germination potential.