A conserved microRNA module exerts homeotic control over Petunia hybrida and Antirrhinum majus floral organ identity

Abstract

It is commonly thought that deep phylogenetic conservation of plant microRNAs (miRNAs) and their targets 1,2 indicates conserved regulatory functions. We show that the blind (bl) mutant of Petunia hybrida 3 and the fistulata (fis) mutant of Antirrhinum majus 4,5 , which have similar homeotic phenotypes, are recessive alleles of two homologous miRNA-encoding genes. The BL and FIS genes control the spatial restriction of homeotic class C genes 6,7 to the inner floral whorls, but their ubiquitous early floral expression patterns are in contradiction with a potential role in patterning C gene expression. We provide genetic evidence for the unexpected function of the MIRFIS and MIRBL genes in the center of the flower and propose a dynamic mechanism underlying their regulatory role. Notably, Arabidopsis thaliana, a more distantly related species, also contains this miRNA module but does not seem to use it to confine early C gene expression to the center of the flower.The spatial partitioning of floral homeotic gene expression is crucial for wild-type flower development. Several transcription factors participate in this control, which aims at transcriptional silencing of the so-called 'C genes' outside their genuine expression domain in the inner two whorls of the flower, where they govern reproductive organ (stamen and carpel) development 6,7 . The functions of orthologous repressor genes, constituting the A function of the floral ABCs 6 , are, in part, comparable between different species 8 , as are some of the cis-acting regulatory regions within the large second intron of their structurally and functionally related target C genes AGAMOUS (AG) in Arabidopsis thaliana 7 , pMADS3 in P. hybrida 9 and PLENA and FARINELLI (PLE and FAR) in A. majus 10 . There are also exceptions to these overall similarities among species. For instance, orthologs of the A. thaliana APETALA2 (AP2) gene have no role in C gene regulation in P. hybrida 11 or A. majus 12 , raising the question of whether other genes fulfill this role. Candidates are the BL gene in P. hybrida and FIS in A. majus, which, when mutated, show markedly similar homeotically converted stamenoid petals in their second floral whorls 4,5 (Fig. 1).By a combination of transposon tagging and map-based cloning strategies, we cloned the BL and FIS genes and found that they encode homologous bona fide miRNAs (miRBL and miRFIS), related in their core sequences to members of the large miR169 family 13,14 (Fig. 2). The bl-1 and fis-1 alleles lie within large genomic deletions and thus represent null alleles; bl-2 and fis-2 are transposon induced and genetically unstable alleles (Fig. 2a). miRNA-encoding genes are relatively small targets for mutation, and therefore, recessive mutants are infrequent; bl and fis thus offer a rare opportunity to study and compare the function of potential orthologs in two plant species. miRNAs control gene expression by recognizing short complementary sequences in their transcripts (miRNA-recognition elements, or MREs), which are then post-transcrip...

Dual Effects of miR156-TargetedSPLGenes andCYP78A5/KLUHon Plastochron Length and Organ Size inArabidopsis thaliana

“…In se-1 and ago1-27 mutants as well as Pro35S:MIM156 plants, all of which have reduced miR156 activity, we observed a clear increase in the levels of the SPL9 target mRNA but no obvious expansion of its expression domain. Thus, miR156 appears to be at the other end of possibilities for miRNA target interactions, with its main role being quantitative control of target mRNA levels, similar to what has been reported for miR169 and its targets in snapdragon (Antirrhinum majus) and petunia (Petunia hybrida; Cartolano et al, 2007). This conclusion is supported by a comparison of SPL9 and miR156 expression patterns, which strongly overlap (Figure 3).…”

Mutagenesis in Petunia x hybrida Vilm. and isolation of a novel morphological mutant

“…Recently, with the completion of several genome and EST projects, the amount of sequence data has increased drastically and allowed the effective use of mutant populations to investigate gene function in high-throughput reverse genetic approaches (Alonso and Ecker, 2006). In petunia, gene function studies have profited from the endogenous system of transposable elements (van den Broeck et al, 1998) to characterize the genetic determinants of a wide range of biological processes (Baumann et al, 2007;Cartolano et al, 2007;Simons et al, 2007). Although highly effective, transposon tagging employing non-engineered sequences can only generate loss-of-function mutations and the unstable nature of the insertion may impair long-term genetic analyses (Alonso and Ecker, 2006).…”

Redefining C and D in the Petunia ABC

“…Furthermore, to date, no A-function could be attributed to petunia orthologs of the Arabidopsis A-function genes. Instead, we found that in both petunia and snapdragon one aspect of the A-function, restriction of C-class activity to stamens and carpels, is encoded by a microRNA, named BLIND in petunia and FISTULATA in snapdragon (Antirrhinum majus; Cartolano et al, 2007).…”