Evolution of the miR5200-FLOWERING LOCUS T flowering time regulon in the temperate grass subfamily Pooideae.

Flowering time is a carefully regulated trait controlled primarily through the action of the central genetic regulator, FLOWERING LOCUS T (FT). Recently it was demonstrated that a microRNA, miR5200, targets the end of the second exon of FT under short-day photoperiods in the grass subfamily Pooideae, thus preventing FT transcripts from reaching threshold levels under non-inductive conditions. Pooideae are an interesting group in that they rapidly diversified from the tropics into the northern temperate region during a major global cooling event spanning the Eocene-Oligocene transition. We hypothesize that miR5200 photoperiod-sensitive regulation of Pooideae flowering time networks assisted their transition into northern seasonal environments. Here, we test predictions derived from this hypothesis that miR5200, originally found in bread wheat and later identified in Brachypodium distachyon, (1) was present in the genome of the Pooideae common ancestor, (2) is transcriptionally regulated by photoperiod, and (3) is negatively correlated with FT transcript abundance, indicative of miR5200 regulating FT. Our results demonstrate that miR5200 did evolve at or around the base of Pooideae, but only acquired photoperiod-regulated transcription within the Brachypodium lineage. Based on expression profiles and previous data, we posit that the progenitor of miR5200 was co-regulated with FT by an unknown mechanism.

Evidence for an Early Origin of Vernalization Responsiveness in Temperate Pooideae Grasses.

The ability of plants to match their reproductive output with favorable environmental conditions has major consequences both for lifetime fitness and geographic patterns of diversity. In temperate ecosystems, some plant species have evolved the ability to use winter nonfreezing cold (vernalization) as a cue to ready them for spring flowering. However, it is unknown how important the evolution of vernalization responsiveness has been for the colonization and subsequent diversification of taxa within the northern and southern temperate zones. Grasses of subfamily Pooideae, including several important crops, such as wheat (Triticum aestivum), barley (Hordeum vulgare), and oats (Avena sativa), predominate in the northern temperate zone, and it is hypothesized that their radiation was facilitated by the early evolution of vernalization responsiveness. Predictions of this early origin hypothesis are that a response to vernalization is widespread within the subfamily and that the genetic basis of this trait is conserved. To test these predictions, we determined and reconstructed vernalization responsiveness across Pooideae and compared expression of wheat vernalization gene orthologs VERNALIZATION1 (VRN1) and VRN3 in phylogenetically representative taxa under cold and control conditions. Our results demonstrate that vernalization responsive Pooideae species are widespread, suggesting that this trait evolved early in the lineage and that at least part of the vernalization gene network is conserved throughout the subfamily. These results are consistent with the hypothesis that the evolution of vernalization responsiveness was important for the initial transition of Pooideae out of the tropics and into the temperate zone.

Comparative Transcriptomics Indicates a Role for SHORT VEGETATIVE PHASE (SVP) Genes in Mimulus guttatus Vernalization Response.

The timing of reproduction in response to variable environmental conditions is critical to plant fitness, and is a major driver of taxon differentiation. In the yellow monkey flower, Mimulus guttatus, geographically distinct North American populations vary in their photoperiod and chilling (vernalization) requirements for flowering, suggesting strong local adaptation to their surroundings. Previous analyses revealed quantitative trait loci (QTL) underlying short-day mediated vernalization responsiveness using two annual M. guttatus populations that differed in their vernalization response. To narrow down candidate genes responsible for this variation, and to reveal potential downstream genes, we conducted comparative transcriptomics and quantitative PCR (qPCR) in shoot apices of parental vernalization responsive IM62, and unresponsive LMC24 inbred lines grown under different photoperiods and temperatures. Our study identified several metabolic, hormone signaling, photosynthetic, stress response, and flowering time genes that are differentially expressed between treatments, suggesting a role for their protein products in short-day-mediated vernalization responsiveness. Only a small subset of these genes intersected with candidate genes from the previous QTL study, and, of the main candidates tested with qPCR under nonpermissive conditions, only SHORT VEGETATIVE PHASE (SVP) gene expression met predictions for a population-specific short-day-repressor of flowering that is repressed by cold.

Evolution of VRN2/Ghd7-Like Genes in Vernalization-Mediated Repression of Grass Flowering.

Flowering of many plant species is coordinated with seasonal environmental cues such as temperature and photoperiod. Vernalization provides competence to flower after prolonged cold exposure, and a vernalization requirement prevents flowering from occurring prior to winter. In winter wheat (Triticum aestivum) and barley (Hordeum vulgare), three genes VRN1, VRN2, and FT form a regulatory loop that regulates the initiation of flowering. Prior to cold exposure, VRN2 represses FT. During cold, VRN1 expression increases, resulting in the repression of VRN2, which in turn allows activation of FT during long days to induce flowering. Here, we test whether the circuitry of this regulatory loop is conserved across Pooideae, consistent with their niche transition from the tropics to the temperate zone. Our phylogenetic analyses of VRN2-like genes reveal a duplication event occurred before the diversification of the grasses that gave rise to a CO9 and VRN2/Ghd7 clade and support orthology between wheat/barley VRN2 and rice (Oryza sativa) Ghd7 Our Brachypodium distachyon VRN1 and VRN2 knockdown and overexpression experiments demonstrate functional conservation of grass VRN1 and VRN2 in the promotion and repression of flowering, respectively. However, expression analyses in a range of pooids demonstrate that the cold repression of VRN2 is unique to core Pooideae such as wheat and barley. Furthermore, VRN1 knockdown in B. distachyon demonstrates that the VRN1-mediated suppression of VRN2 is not conserved. Thus, the VRN1-VRN2 feature of the regulatory loop appears to have evolved late in the diversification of temperate grasses.

Phylogenetic analyses place the Australian monotypic Revwattsia in Dryopteris (Dryopteridaceae).

Revwattsia fragilis (Watts) D.L. Jones (Dryopteridaceae), originally described as a Polystichum Roth by the pioneer Australian botanist Reverend W.W. Watts in 1914, is a rare epiphytic fern endemic to northeastern Queensland, Australia. Known from only a few populations, it is restricted to tropical rainforests in the Atherton Tablelands. We used the cpDNA markers psbA-trnH, rbcL, rbcL-accD, rps4-trnS, trnG-trnR, trnL-trnF, and trnP-petG to infer the relationships of Revwattsia fragilis within Dryopteridaceae. Based on our molecular analysis, we were able to reject Watts's 1914 hypothesis of a close relationship to Polystichum. Its closest allies are a suite of Asian Dryopteris Adans. species including Dryopteris labordei, Dryopteris gymnosora, Dryopteris erythrosora and Dryopteris cystolepidota; maintaining Revwattsia renders Dryopteris paraphyletic. The epiphytic habit and distinctive long-creeping rhizome of Revwattsia appear to be autapomorphies and do not warrant its generic status. In the course of our investigation we confirmed that polyphyly of Dryopteris is also sustained by the inclusion of Acrorumohra (H.Itô) H.Itô, Acrophorus C.Presl, Arachniodes Blume, Diacalpe Blume, Dryopsis Holttum & P.J.Edwards, and Peranema D.Don. The epithet fragilis is occupied in Dryopteris, therefore we provide the name Dryopteris wattsiinom. nov. to accommodate Revwattsia fragilis in Dryopteris.