DEF- and GLO-like proteins may have lost most of their interaction partners during angiosperm evolution.

BACKGROUND AND AIMS DEFICIENS (DEF)- and GLOBOSA (GLO)-like proteins constitute two sister clades of floral homeotic transcription factors that were already present in the most recent common ancestor (MRCA) of extant angiosperms. Together they specify the identity of petals and stamens in flowering plants. In core eudicots, DEF- and GLO-like proteins are functional in the cell only as heterodimers with each other. There is evidence that this obligate heterodimerization contributed to the canalization of the flower structure of core eudicots during evolution. It remains unknown as to whether this strict heterodimerization is an ancient feature that can be traced back to the MRCA of extant flowering plants or if it evolved later during the evolution of the crown group angiosperms. METHODS The interactions of DEF- and GLO-like proteins of the early-diverging angiosperms Amborella trichopoda and Nuphar advena and of the magnoliid Liriodendron tulipifera were analysed by employing yeast two-hybrid analysis and electrophoretic mobility shift assay (EMSA). Character-state reconstruction, including data from other species as well, was used to infer the ancestral interaction patterns of DEF- and GLO-like proteins. KEY RESULTS The yeast two-hybrid and EMSA data suggest that DEF- and GLO-like proteins from early-diverging angiosperms both homo- and heterodimerize. Character-state reconstruction suggests that the ability to form heterodimeric complexes already existed in the MRCA of extant angiosperms and that this property remained highly conserved throughout angiosperm evolution. Homodimerization of DEF- and GLO-like proteins also existed in the MRCA of all extant angiosperms. DEF-like protein homodimerization was probably lost very early in angiosperm evolution and was not present in the MRCA of eudicots and monocots. GLO-like protein homodimerization might have been lost later during evolution, but very probably was not present in the MRCA of eudicots. CONCLUSIONS The flexibility of DEF- and GLO-like protein interactions in early-diverging angiosperms may be one reason for the highly diverse flower morphologies observed in these species. The results strengthen the hypothesis that a reduction in the number of interaction partners of DEF- and GLO-like proteins, with DEF-GLO heterodimers remaining the only DNA-binding dimers in core eudicots, contributed to developmental robustness, canalization of flower development and the diversification of angiosperms.