The identity of Antirrhinum reproductive organs is controlled by two class C- MADS box genes, PLENA (PLE) and FARINELLI (FAR), which are expressed in the third and the fourth whorl of the flower. The fistulata mutant (fis) in Antirrhinum and blind (bl) in Petunia show partial homeotic conversion of petals to stamenoid structures, which is caused by ectopic expression of PLE/FAR and pMADS3/FBP6, the Petunia C-genes, respectively. FIS and BL thus control the C- expression domain in the wild type. FISTULATA was cloned by a map-based strategy combined with a candidate gene approach and show that both genes encode a microRNA corresponding to miR169. The miR169 target site is present in the 3´UTR of members of the NF-YA transcription factor family that bind a cis-acting CCAAT motif present in the second intron of the PLE/FAR and pMADS3 genes. This suggests a miRNA-mediated and NF-YA-dependent control of the C- function. We cloned six members of the Antirrhinum NF-YA gene family (AmYAs), four of which contain the miRNA recognition site. Data derived from in vitro infitration experiments indicate miRFIS-dependent post-transcriptional regulation of AmYAs. Expression studies in wild type and fis mutant flowers showed that miRFIS and the AmYAs are spatially co-expressed in the flower suggesting that by fine tuning expression of the NF-YAs, miRFIS indirectly regulates the level of C-gene expression. In addition, genetic evidence is presented demonstrating the functional relevance of the unexpected miRFIS function in the centre of the flower. Based on these results a dynamic model is proposed in which miRFIS maintains the boundaries of the C- domain by preventing formation of a radially extending molecular gradient from the centre of the flower (where C-products are activated) toward peripheral regions of the meristem. Experimental approaches are suggested to solve open questions
