Beyond cyanogenesis: Temperature gradients drive environmental adaptation in North American white clover (Trifolium repens L.).

Species that repeatedly evolve phenotypic clines across environmental gradients have been highlighted as ideal systems for characterizing the genomic basis of local environmental adaptation. However, few studies have assessed the importance of observed phenotypic clines for local adaptation: conspicuous traits that vary clinally may not necessarily be the most critical in determining local fitness. The present study was designed to fill this gap, using a plant species characterized by repeatedly evolved adaptive phenotypic clines. White clover is naturally polymorphic for its chemical defence cyanogenesis (HCN release with tissue damage); climate-associated cyanogenesis clines have evolved throughout its native and introduced range worldwide. We performed landscape genomic analyses on 415 wild genotypes from 43 locations spanning much of the North American species range to assess the relative importance of cyanogenesis loci vs. other genomic factors in local climatic adaptation. We find clear evidence of local adaptation, with temperature-related climatic variables best describing genome-wide differentiation between sampling locations. The same climatic variables are also strongly correlated with cyanogenesis frequencies and gene copy number variations (CNVs) at cyanogenesis loci. However, landscape genomic analyses indicate no significant contribution of cyanogenesis loci to local adaptation. Instead, several genomic regions containing promising candidate genes for plant response to seasonal cues are identified - some of which are shared with previously identified QTLs for locally adaptive fitness traits in North American white clover. Our findings suggest that local adaptation in white clover is likely determined primarily by genes controlling the timing of growth and flowering in response to local seasonal cues. More generally, this work suggests that caution is warranted when considering the importance of conspicuous phenotypic clines as primary determinants of local adaptation.

Genetic trade-offs underlie divergent life history strategies for local adaptation in white clover.

Local adaptation is common in plants, yet characterization of its underlying genetic basis is rare in herbaceous perennials. Moreover, while many plant species exhibit intraspecific chemical defence polymorphisms, their importance for local adaptation remains poorly understood. We examined the genetic architecture of local adaptation in a perennial, obligately-outcrossing herbaceous legume, white clover (Trifolium repens). This widespread species displays a well-studied chemical defence polymorphism for cyanogenesis (HCN release following tissue damage) and has evolved climate-associated cyanogenesis clines throughout its range. Two biparental F2  mapping populations, derived from three parents collected in environments spanning the U.S. latitudinal species range (Duluth, MN, St. Louis, MO and Gainesville, FL), were grown in triplicate for two years in reciprocal common garden experiments in the parental environments (6,012 total plants). Vegetative growth and reproductive fitness traits displayed trade-offs across reciprocal environments, indicating local adaptation. Genetic mapping of fitness traits revealed a genetic architecture characterized by allelic trade-offs between environments, with 100% and 80% of fitness QTL in the two mapping populations showing significant QTL×E interactions, consistent with antagonistic pleiotropy. Across the genome there were three hotspots of QTL colocalization. Unexpectedly, we found little evidence that the cyanogenesis polymorphism contributes to local adaptation. Instead, divergent life history strategies in reciprocal environments were major fitness determinants: selection favoured early investment in flowering at the cost of multiyear survival in the southernmost site versus delayed flowering and multiyear persistence in the northern environments. Our findings demonstrate that multilocus genetic trade-offs contribute to contrasting life history characteristics that allow for local adaptation in this outcrossing herbaceous perennial.

Intraspecific variation in elemental accumulation and its association with salt tolerance in Paspalum vaginatum.

Most plant species, including most crops, perform poorly in salt-affected soils because high sodium levels are cytotoxic and can disrupt the uptake of water and important nutrients. Halophytes are species that have evolved adaptations to overcome these challenges and may be a useful source of knowledge for salt tolerance mechanisms and genes that may be transferable to crop species. The salt content of saline habitats can vary dramatically by location, providing ample opportunity for different populations of halophytic species to adapt to their local salt concentrations; however, the extent of this variation, and the physiology and polymorphisms that drive it, remain poorly understood. Differential accumulation of inorganic elements between genotypes or populations may play an important role in local salinity adaptation. To test this, we investigated the relationships between population structure, tissue ion concentrations, and salt tolerance in 17 "fine-textured" genotypes of the halophytic turfgrass seashore paspalum (Paspalum vaginatum Swartz). A high-throughput ionomics pipeline was used to quantify the shoot concentration of 18 inorganic elements across three salinity treatments. We found a significant relationship between population structure and ion accumulation, with strong correlations between principal components derived from genetic and ionomic data. Additionally, genotypes with higher salt tolerance accumulated more K and Fe and less Ca than less tolerant genotypes. Together these results indicate that differences in ion accumulation between P. vaginatum populations may reflect locally adapted salt stress responses.

Dual-species origin of an adaptive chemical defense polymorphism.

Allopolyploid speciation and chemical defense diversification are two of the most characteristic features of plant evolution; although the former has likely shaped the latter, this has rarely been documented. Here we document allopolyploidy-mediated chemical defense evolution in the origin of cyanogenesis (HCN release upon tissue damage) in white clover (Trifolium repens). We combined linkage mapping of the loci that control cyanogenesis (Ac, controlling production of cyanogenic glucosides; and Li, controlling production of their hydrolyzing enzyme linamarase) with genome sequence comparisons between white clover, a recently evolved allotetraploid, and its diploid progenitors (Trifolium pallescens, Trifolium occidentale). The Ac locus (a three-gene cluster comprising the cyanogenic glucoside pathway) is derived from T. occidentale; it maps to linkage group 2O (occidentale subgenome) and is orthologous to a highly similar cluster in the T. occidentale reference genome. By contrast, Li maps to linkage group 4P (pallescens subgenome), indicating an origin in the other progenitor species. These results indicate that cyanogenesis evolved in white clover as a product of the interspecific hybridization that created the species. This allopolyploidization-derived chemical defense, together with subsequent selection on intraspecific cyanogenesis variation, appears to have contributed to white clover's ecological success as a globally distributed weed species.

Hybridization, polyploidy and clonality influence geographic patterns of diversity and salt tolerance in the model halophyte seashore paspalum (Paspalum vaginatum).

In plant species, variation in levels of clonality, ploidy and interspecific hybridization can interact to influence geographic patterns of genetic diversity. These factors commonly vary in plants that specialize on saline habitats (halophytes) and may play a role in how they adapt to salinity variation across their range. One such halophyte is the turfgrass and emerging genomic model system seashore paspalum (Paspalum vaginatum Swartz). To investigate how clonal propagation, ploidy variation, and interspecific hybridization vary across ecotypes and local salinity levels in wild P. vaginatum, we employed genotyping-by-sequencing, cpDNA sequencing and flow cytometry in 218 accessions representing > 170 wild collections from throughout the coastal southern United States plus USDA germplasm. We found that the two morphologically distinct ecotypes of P. vaginatum differ in their adaptive strategies. The fine-textured ecotype is diploid and appears to reproduce in the wild both sexually and by clonal propagation; in contrast, the coarse-textured ecotype consists largely of clonally-propagating triploid and diploid genotypes. The coarse-textured ecotype appears to be derived from hybridization between fine-textured P. vaginatum and an unidentified Paspalum species. These clonally propagating hybrid genotypes are more broadly distributed than clonal fine-textured genotypes and may represent a transition to a more generalist adaptive strategy. Additionally, the triploid genotypes vary in whether they carry one or two copies of the P. vaginatum subgenome, indicating multiple evolutionary origins. This variation in subgenome composition shows associations with local ocean salinity levels across the sampled populations and may play a role in local adaptation.

Sterile Spikelets Contribute to Yield in Sorghum and Related Grasses.

Sorghum (Sorghum bicolor) and its relatives in the grass tribe Andropogoneae bear their flowers in pairs of spikelets in which one spikelet (seed-bearing or sessile spikelet [SS]) of the pair produces a seed and the other is sterile or male (staminate). This division of function does not occur in other major cereals such as wheat (Triticum aestivum) or rice (Oryza sativa). Additionally, one bract of the SS spikelet often produces a long extension, the awn, that is in the same position as, but independently derived from, that of wheat and rice. The function of the sterile spikelet is unknown and that of the awn has not been tested in Andropogoneae. We used radioactive and stable isotopes of carbon, RNA sequencing of metabolically important enzymes, and immunolocalization of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) to show that the sterile spikelet assimilates carbon, which is translocated to the largely heterotrophic SS. The awn shows no evidence of photosynthesis. These results apply to distantly related species of Andropogoneae. Removal of sterile spikelets in sorghum significantly decreases seed weight (yield) by ∼9%. Thus, the sterile spikelet, but not the awn, affects yield in the cultivated species and fitness in the wild species.

Identification of Novel QTL Conferring Sheath Blight Resistance in Two Weedy Rice Mapping Populations.

BACKGROUND: Rice sheath blight (ShB) disease, caused by the pathogenic fungus Rhizoctonia solani, causes significant yield losses globally. US weedy rice populations, which are de-domesticated forms of indica and aus cultivated rice, appear to be more resistant to ShB than local japonica cultivated rice. We mapped quantitative trait loci (QTL) associated with ShB resistance using two F8 recombinant inbred line populations generated from crosses of an indica crop variety, Dee-Geo-Woo-Gen (DGWG), with individuals representing the two major US weed biotypes, straw hull (SH) and black hull awned (BHA). RESULTS: We identified nine ShB resistance QTL across both mapping populations. Five were attributable to alleles that affect plant height (PH) and heading date (HD), two growth traits that are known to be highly correlated with ShB resistance. By utilizing an approach that treated growth traits as covariates in the mapping model, we were able to infer that the remaining four QTL are involved in ShB resistance. Two of these, qShB1-2 and qShB4, are different from previously identified ShB QTL and represent new candidates for further study. CONCLUSION: Our findings suggest that ShB resistance can be improved through favorable plant growth traits and the combined effects of small to moderate-effect resistance QTL. Additionally, we show that including PH and HD as covariates in QTL mapping models is a powerful way to identify new ShB resistance QTL.

Comprehensive identification and clustering of CLV3/ESR-related (CLE) genes in plants finds groups with potentially shared function.

CLV3/ESR (CLE) proteins are important signaling peptides in plants. The short CLE peptide (12-13 amino acids) is cleaved from a larger pre-propeptide and functions as an extracellular ligand. The CLE family is large and has resisted attempts at classification because the CLE domain is too short for reliable phylogenetic analysis and the pre-propeptide is too variable. We used a model-based search for CLE domains from 57 plant genomes and used the entire pre-propeptide for comprehensive clustering analysis. In total, 1628 CLE genes were identified in land plants, with none recognizable from green algae. These CLEs form 12 groups within which CLE domains are largely conserved and pre-propeptides can be aligned. Most clusters contain sequences from monocots, eudicots and Amborella trichopoda, with sequences from Picea abies, Selaginella moellendorffii and Physcomitrella patens scattered in some clusters. We easily identified previously known clusters involved in vascular differentiation and nodulation. In addition, we found a number of discrete groups whose function remains poorly characterized. Available data indicate that CLE proteins within a cluster are likely to share function, whereas those from different clusters play at least partially different roles. Our analysis provides a foundation for future evolutionary and functional studies.