Gene Duplication and Transference of Function in the paleo Lineage of Floral Organ Identity Genes.

The floral organ identity gene () is a MADS-box transcription factor involved in stamen and petal identity that belongs to the B-class of the ABC model of flower development. (Ranunculaceae), an emerging model in the non-core eudicots, has homologs derived from both ancient and recent gene duplications. Prior work has shown that petals have been lost repeatedly and independently in Ranunculaceae in correlation with the loss of a specific paralog, and represents one of these instances. The main goal of this study was to conduct a functional analysis of the three orthologs present in , representing the paleo gene lineage, to determine the degree of redundancy versus divergence after gene duplication. Because lacks petals, and has lost the petal-specific , we also asked whether heterotopic expression of the remaining genes contributes to the partial transference of petal function to the first whorl found in insect-pollinated species. To address these questions, we undertook functional characterization by virus-induced gene silencing (VIGS), protein-protein interaction and binding site analyses. Our results illustrate partial redundancy among s, with deep conservation of B-class function in stamen identity and a novel role in ectopic petaloidy of sepals. Certain aspects of petal function of the lost locus have apparently been transferred to the other paralogs. A novel result is that the protein products interact not only with each other, but also as homodimers. Evidence presented here also suggests that expression of the different paralogs is tightly integrated, with an apparent disruption of B function homeostasis upon silencing of one of the paralogs that codes for a truncated protein. To explain this result, we propose two testable alternative scenarios: that the truncated protein is a dominant negative mutant or that there is a compensational response as part of a back-up circuit. The evidence for promiscuous protein-protein interactions via yeast two-hybrid combined with the detection of AP3 specific binding motifs in all B-class gene promoters provide partial support for these hypotheses.