Developmental genetics of corolla tube formation in monkeyflowers (Mimulus): role of the tasiRNA-ARF pathway

About one third of all angiosperm species produce flowers with petals fused into a corolla tube. As an important element of the tremendous diversity of flower morphology, the corolla tube plays a critical role in many specialized interactions between plants and animal pollinators (e.g., beeflies, hawkmoths, hummingbirds, nectar bats), which in turn drives rapid plant speciation. Despite its clear significance in plant reproduction and evolution, the corolla tube remains one of the least understood plant structures from a developmental genetics perspective. Through mutant analyses and transgenic experiments, here we show that the tasiRNA-ARF pathway is required for corolla tube formation in the monkeyflower species Mimulus lewisii. Loss-of-function mutations in the M. lewisii orthologs of ARGONAUTE7 and SUPRESSOR OF GENE SILENCING 3 cause a dramatic decrease in tasiARF abundance and a moderate up-regulation of Auxin Response Factor 3 (ARF3) and ARF4, which lead to inhibition of lateral expansion of the bases of petal primordia and complete arrest of the upward growth of the inter-primordial regions, resulting in unfused corollas. Integrating our molecular and phenotypic analyses of the tasiRNA-ARF pathway in Mimulus with historical insights from morphological and anatomical studies in various sympetalous species, we propose a new conceptual model for the developmental genetic control of corolla tube formation. Overall design: The goal of this experiment is to determine tasiRNA abundance in the Mimulus lewisii mutants flayed1 and flayed2 compared to the wild-type (inbred line LF10). The six samples include two biological replicates of the wild-type LF10 (LF10A and LF10B), two biological replicates of the flayed1 mutant (flayed1A and flayed1B), and two biological replicates of the flayed2 mutant (flayed2A and flayed2B). All plants were grown under the same standard greenhouse environment (22 oC; light:dark = 16h:8h). For all samples, RNA was extracted from the same tissue at the same developmental stage (i.e., 5-mm floral buds)