Evolution of the basic Helix-Loop-Helix transcription factor SPATULA and its role in gynoecium development.

Abstract

SPATULA (SPT) encodes a basic Helix-Loop-Helix transcription factor in Arabidopsis thaliana that functions in the development of the style, stigma and replum tissues, all of which arise from the carpel margin meristem (CMM) of the gynoecium. Here, we use a comparative approach to investigate the evolutionary history of SPT and identify changes that potentially contributed to its role in gynoecium development. We investigate SPT's molecular and functional evolution using phylogenetic reconstruction, yeast-2-hybrid analyses of protein-protein interactions, microarray-based analyses of protein-DNA interactions, plant transformation assays, RNA in-situ hybridization, and in-silico analyses of promoter sequences. We demonstrate the SPT lineage to have arisen at the base of euphyllophytes from a clade of potentially light-regulated transcription factors through gene duplication followed by the loss of an Active Phytochrome Binding (APB) domain. We also clarify the more recent evolutionary history of SPT and its paralog ALCATRAZ (ALC), which appear to have arisen through a large-scale duplication within Brassicales. We find that SPT orthologs from diverse groups of seed plants share strikingly similar capacities for protein-protein and protein-DNA interactions, and that SPT coding regions from a wide taxonomic range of plants are able to complement loss-of-function spt mutations in transgenic Arabidopsis. However, the expression pattern of SPT appears to have evolved significantly within angiosperms, and we identify structural changes in SPT's promoter region that correlate with the acquisition of high expression levels in tissues arising from the CMM in Brassicaeae. We conclude that changes to SPT's expression pattern made a major contribution to the evolution of its developmental role in the gynoecium of Brassicaeae. By contrast, the main biochemical capacities of SPT, as well as many of its immediate transcriptional targets, appear to have been conserved at least since the base of living angiosperms.