Panicum represents a large genus of many North American prairie grass species. These include switchgrass (Panicum virgatum), a biofuel crop candidate with wide geographic range, as well as Panicum hallii, a close relative to switchgrass, which serves as a model system for the study of Panicum genetics due to its diploid genome and short growth cycles. For the advancement of switchgrass as a biofuel crop, it is essential to understand host microbiome interactions, which can be impacted by plant genetics and environmental factors inducing ecotype-specific phenotypic traits. We here compared rhizosphere and root endosphere bacterial communities of upland and lowland P. virgatum and P. hallii genotypes planted at two sites in Texas. Our analysis shows that sampling site predominantly contributed to bacterial community variance in the rhizosphere, however, impacted root endosphere bacterial communities much less. Instead we observed a relatively large core endophytic microbiome dominated by ubiquitously root-colonizing bacterial genera Streptomyces, Pseudomonas, and Bradyrhizobium. Endosphere communities displayed comparable diversity and conserved community structures across genotypes of both Panicum species. Functional insights into interactions between P. hallii and its root endophyte microbiome could hence inform testable hypotheses that are relevant for the improvement of switchgrass as a biofuel crop.

Local adaptation is the process by which natural selection drives adaptive phenotypic divergence across environmental gradients. Theory suggests that local adaptation results from genetic tradeoffs at individual genetic loci, where adaptation to one set of environmental conditions results in a cost to fitness in alternative environments. However, the degree to which there are costs associated with local adaptation is poorly understood because most of these experiments rely on two-site reciprocal transplant experiments. Here, we quantify the benefits and costs of locally adaptive loci across 17° of latitude in a four-grandparent outbred mapping population in outcrossing switchgrass (Panicum virgatum L.), an emerging biofuel crop and dominant tallgrass species. We conducted quantitative trait locus (QTL) mapping across 10 sites, ranging from Texas to South Dakota. This analysis revealed that beneficial biomass (fitness) QTL generally incur minimal costs when transplanted to other field sites distributed over a large climatic gradient over the 2 y of our study. Therefore, locally advantageous alleles could potentially be combined across multiple loci through breeding to create high-yielding regionally adapted cultivars.

The rice landrace Horkuch, endemic to the southern saline coast of Bangladesh, is known to have salt tolerance traits and can therefore contribute to a high yielding recipient for breeding purposes. In this study, we reciprocally crossed Horkuch with high yielding but salt sensitive IR29 to detect the complement of genes that were responsible for conferring salt tolerance versus sensitivity at the seedling developmental stage. We looked at tolerant and sensitive F₃ families from individual F₂ segregating plants and analyzed them for differential gene expressions using RNAseq. In general, we observed higher numbers of genes differentially expressed in leaves compared to root tissues. This included both upregulation and downregulation of gene expression across our experimental factors. Gene expression decreased in sensitive leaf after stress exposure where tolerant plants showed the opposite trend. In root, tolerant plants expression decreased at higher time points of stress exposure. We also observed a strong maternal cytoplasmic effect on gene expression and this was most evident in roots where there was upregulation in functional enrichments related to phosphorylation, electron carriers, transporter and cation transmembrane activities. Stress groups (tolerant and sensitive) response in F₃ families were distinctive in both cytoplasmic backgrounds and involved uniquely upregulated genes in tolerant progenies including membrane sensor proteins, enzymes involved with signaling pathways, such as those producing trehalose and G-protein coupled receptor proteins, photosynthesis-related enzymes and golgi body recycling as well as prolamin precursor proteins involved in refolding of proteins. On the other hand, sensitivity was found to be associated with differential upregulation of only a few redox proteins and higher number of apoptosis related genes compared to the tolerant response. Overall, our highly replicated experimental design was powerful and allowed the detection of relatively subtle differential expression. Our future goal is to correlate these expression differences with QTLs in this population, which would help identify the relative importance of specific genetic loci and provide a direct avenue for combining higher levels of salt tolerance with better agronomic traits in rice.

The success or failure of reproductive outcomes from intra-species crosses between plants of diferent ploidy levels is an important factor in fowering plant evolution and crop breeding. However, the efects of inter-ploidy cross directions on F1 hybrid ofspring ftness are poorly understood. In Arabidopsis thaliana, hybridization between diploid and tetraploid plants can produce viable F1 triploid plants. When selfed, such F1 triploid plants act as aneuploid gamete production “machines” where the vast majority of gametes generated are aneuploid which, following sexual reproduction, can generate aneuploid swarms of F2 progeny (Henry et al. 2009). There is potential for some aneuploids to cause gametophyte abortion and/or F2 seed abortion (Henry et al. 2009). In this study, we analyse the reproductive success of 178 self-fertilized inter-accession F1 hybrid triploids and demonstrate that the proportions of aborted or normally developed F2 seeds from the selfed F1 triploids depend upon a combination of natural variation and cross direction, with strong interaction between these factors. Singleseed ploidy analysis indicates that the embryonic DNA content of phenotypically normal F2 seeds is highly variable and that these DNA content distributions are also afected by genotype and cross direction. Notably, genetically identical reciprocal F1 hybrid triploids display grandparent-of-origin efects on F2 seed set, and hence on the ability to tolerate aneuploidy in F2 seed. There are diferences between reciprocal F1 hybrid triploids regarding the proportions of normal and aborted F2 seeds generated, and also for the DNA content averages and distributions of the F2 seeds. To identify genetic variation for tolerance of aneuploidy in F2 seeds, we carried out a GWAS which identifed two SNPs, termed MOT and POT, which represent candidate loci for genetic control of the proportion of normal F2 seeds obtained from selfed F1 triploids. Parental and grandparental efects on F2 seeds obtained from selfed F1 triploids can have transgenerational consequences for asymmetric gene fow, emergence of novel genotypes in polyploid populations, and for control of F2 seed set in triploid crops.

Environmental heterogeneity can drive patterns of functional trait variation and lead to the formation of locally adapted ecotypes. Plant ecotypes are often differentiated by suites of correlated root and shoot traits that share common genetic, developmental, and physiological relationships. For instance, although plant water loss is largely governed by shoot systems, root systems determine water access and constrain shoot water status. To evaluate the genetic basis of root and shoot trait divergence, we developed a recombinant inbred population derived from mesic and xeric ecotypes of the perennial grass Panicum hallii. Our study sheds light on the genetic architecture underlying the relationships between root and shoot traits. We identified several genomic ‘hotspots’ which control suites of correlated root and shoot traits, thus indicating genetic coordination between plant organ systems in the process of ecotypic divergence. Genomic regions of colocalized quantitative trait locus (QTL) for the majority of shoot and root growth related traits were independent of colocalized QTL for shoot and root resource acquisition traits. The allelic effects of individual QTL underscore ecological specialization for drought adaptation between ecotypes and reveal possible hybrid breakdown through epistatic interactions. These results have implications for understanding the factors constraining or facilitating local adaptation in plants.

Photoperiod is a key environmental cue affecting flowering and biomass traits in plants. Key components of the photoperiodic flowering pathway have been identified in many species, but surprisingly few studies have globally examined the diurnal rhythm of gene expression with changes in day‐length. Using a cost‐effective 3'‐Tag RNA sequencing strategy, we characterize 9,010 photoperiod responsive genes with strict statistical testing across a diurnal time series in the C₄ perennial grass, Panicum hallii. We show that the vast majority of photoperiod responses are driven by complex interactions between day‐length and sampling periods. A fine‐scale contrast analysis at each sampling time revealed a detailed picture of the temporal reprogramming of cis‐regulatory elements and biological processes under short and long day conditions. Phase shift analysis reveals quantitative variation among genes with photoperiod‐dependent diurnal patterns. In addition, we identify three photoperiod enriched transcription factor families with key genes involved in photoperiod flowering regulatory networks. Finally, co‐expression networks analysis of GIGANTEA homolog predicted 1,668 potential coincidence partners, including five well‐known GI‐interacting proteins. Our results not only provide a resource for understanding the mechanisms of photoperiod regulation in perennial grasses but also lays a foundation to increase biomass yield in biofuel crops.

Previous studies have shown that switchgrass has a wide range of genetic variation and that productivity is linked to local adaptation to the location of origin for many cultivars. In this meta -analysis, we compiled and analyzed 900 observations associated with 41 field trials for four switchgrass cultivars (two lowlands , Alamo and Kanlow , and two upland s , Cave -In -Rock and Shelter ) . This extensive dataset and machine learning was used to identify the most influential variables impacting switchgrass productivity , to search for evidence of local adaptation to each cultivar’s location of origin, and to predict change in productivity under future climate for each cultivar. In general, variables associated with climate and management are more important predictors of productivity relative to soil variables. Three climatic variables, annual mean temperature , annual precipitation, and precipitation in the wettest month, are identified as key environmental variables for productivity of all cultivars . Productivity under future climate (2041 -2060) is predicted to stay stable for all cultivars relative to the prediction under current climate (1986 -2005) across all trial locations and over a 20 -yr simulation period. However, the productivity of each cultivar varies from location to location and from year to year, although productivity varies more between locations than between years. Additionally, we observe shifts in the most productive cultivar at the local field scale depending on the combination of management practice and climates . The shape of the relationship between productivity and the annual mean temperature relative to the cultivar’s location of origin is bell -shape curve for Kanlow, Cave -in -Rock, and Shelter, indicative of local adaptation. Identifying influential environmental variables , and their relationships to productivity with respect to cultivar’s location of origin helps predicting productivity on the local field scale, and wil l help with the biofuel production planning through the selection of suitable cultivars for different locations under climate changes.

Switchgrass (Panicum virgatum L.) is a perennial C₄ grass native to North America. Over the last two decades switchgrass has received considerable attention for its potential as a bioenergy-biofuel feedstock crop and model system to examine the intraspecific genetic divergence in plants. In April 2017, we observed and collected panicles from the switchgrass accession ‘West Bee Caves’ (WBC3), which was being grown within a four-way mapping population (Milano et al. 2016) planted in 2015 in Temple, TX (UTM: 14 N 657505.8E, 3435580.0N). Infected plants had conspicuously swollen spikelets with reddish- to deep-purple glumes, and, when agitated, spikelets released a dry, red-brown mass of teliospores. Cross-sectioning of spore-laden spikelets revealed the complete replacement of the ovary with fungal sori, consistent with infection by the smut fungus Tilletia maclaganii (Berk.) G.P. Clinton or T. pulcherrima Ellis & Galloway (Tilletiaceae). Likewise, in April 2018 within the same mapping population, Tilletia-infected panicles were collected from WBC3, ‘Alamo’ (AP13), and an F₁-hybrid between ‘Summer’ (VS16) × WBC3 planted in Austin, TX (UTM: 14 N 622078.5E, 3362019.3N), as well as AP13 planted in Kingsville, TX (UTM: 14 N 610479.6E, 3047839.0N). The site in Austin is located ∼100 km south-southwest of Temple, TX, and the site in Kingsville is ∼400 km south of Temple. Seven voucher specimens (n = 3 Austin, 2 Kingsville, and 2 Temple) were deposited in the Cornell Plant Pathology Herbarium (CUP 068303 to 068309). Thereafter, the Tilletia sp. was determined by teliospore morphology as well as polymerase chain reaction (PCR) amplification and sequencing of the internal transcribed spacer (ITS) region using primers ITS5 and ITS4. DNA extracts for PCR were procured from single-teliospore cultures or direct lysis of teliospores from single spikelets. Teliospores (n ≥ 25 per voucher specimen, n = 200 total) lacked a sheath and were pale yellow to orange-brown in color, mostly globose to broadly ellipsoid, rarely irregular, and measured 19.1 to 31.0 × 14.1 to 27.0 µm (mean = 21.5 ± 0.4 × 23.8 ± 0.4 µm). Cell walls were finely verrucose, 1.7 to 3.8 µm thick (mean = 2.7 ± 0.1 µm), and serrate in profile. Teliospore morphology was consistent across host-site combinations and best fit the description of the species T. maclaganii (Vánky 2012). DNA analyses confirmed our morphological-based identification, because ITS sequences were identical across all seven specimens (674 nucleotides; GenBank MH397299 to MH397305) and shared 100% nucleotide identity with authenticated ITS sequences of T. maclaganii from Indiana (GenBank MH256491) and New York State (JF745116; Layton and Bergstrom 2011). To the best of our knowledge, this the first report of the head smut fungus in Texas (Farr and Rossman 2018) and expands the southern geographic range of T. maclaganii in North America to below 28° N latitude. The head smut fungus is an economically important pathogen of switchgrass, particularly when switchgrass is grown across large acreage for biomass, commercial seed, and/or lignocellulosic ethanol (Thomsen et al. 2008). T. maclaganii persists as a systemic parasite within switchgrass plants from which pathogen spread can occur via windborne teliospores, teliospore-infested seed, or asexual propagation of infected plants. Thus, the documentation of T. maclaganii in widespread locations in Texas suggests that management of head smut may become necessary for switchgrass feedstock production in the southern Great Plains.

The stress hormone abscisic acid (ABA) is critical for drought resistance; however, mechanisms controlling ABA levels are unclear. At low water potential, ABA accumulation in the Arabidopsis thaliana accession Shahdara (Sha) was less than that in Landsberg erecta (Ler) or Columbia. Analysis of a Ler x Sha recombinant inbred line population revealed a single major effect quantitative trait locus for ABA accumulation, which included 9-cis-epoxycarotenoid dioxygenase 3 (NCED3) as a candidate gene. NCED3 encodes a rate-limiting enzyme for stress-induced ABA synthesis. Complementation experiments indicated that Sha has a reduced-function NCED3 allele. Compared to Ler, Sha did not have reduced NCED3 gene expression or protein level, but did have four amino acid substitutions within NCED3. Sha differed from Ler in the apparent molecular weight of NCED3, indicative of altered NCED3 proteolytic processing in the chloroplast. Site-directed mutagenesis demonstrated that substitution at amino acid 271 was critical for the altered NCED3 molecular weight pattern while the other Sha NCED3 polymorphisms were also involved in the reduced ABA accumulation. Sha did not have a reduced level of thylakoid-bound NCED3 but did differ from Ler in the apparent molecular weight of stromal NCED3. As Sha was not impaired in known factors critical for NCED3 function in ABA synthesis (expression, chloroplast import, thylakoid binding), the differences between Ler and Sha NCED3 may affect NCED3 activity or other factors influencing NCED3 function. These results identify functionally important sites on NCED3 and indicate a complex pattern of NCED3 post-translational regulation in the chloroplast.

Switchgrass (

Panicum virgatum

) is a promising bio-

fuel crop native to the United States with genotypes

that are adapted to a wide range of distinct ecosys-

tems. Various plants have been shown to undergo

symbioses with plant growth-promoting bacteria and

fungi, however, plant-associated microbial communi-

ties of switchgrass have not been extensively studied

to date. We present 16S ribosomal RNA gene and

internal transcribed spacer (ITS) data of rhizosphere

and root endosphere compartments of four switch-

grass genotypes to test the hypothesis that host

selection of its root microbiota prevails after transfer

to non-native soil. We show that differences in

bacterial, archaeal and fungal community composition

and diversity are strongly driven by plant compart-

ment and switchgrass genotypes and ecotypes.

Plant-associated microbiota show an enrichment in

Alphaproteobacteria

and

Actinobacteria

as well as

Sordariales

and

Pleosporales

compared with the sur-

rounding soil. Root associated compartments display

low-complexity communities dominated and enriched

in

Actinobacteria,

in particular

Streptomyces,

in the

lowland genotypes, and in

Alphaproteobacteria,

speci

fi

cally

Sphingobium

, in the upland genotypes.

Our comprehensive root analysis serves as a snap-

shot of host-speci

fi

c bacterial and fungal associations

of switchgrass in the

fi

eld and con

fi

rms that host-

selected microbiomes persist after transfer to non-

native soil.

Background

Understanding how and why genetic variation is partitioned across geographic space is of fundamental importance to understanding the nature of biological species. How geographical isolation and local adaptation contribute to the formation of ecotypically differentiated groups of plants is just beginning to be understood through population genomic studies. We used whole genome sequencing combined with association study of climate to discover the drivers of differentiation in the perennial C4 grass Panicum hallii.

Results

Sequencing of 89 natural accessions of P.hallii revealed complex population structure across the species range. Major population genomic separation was found between subspecies P.hallii var. hallii and var. filipes as well as between at least four major unrecognized subgroups within var. hallii. At least 139 genomic SNPs were significantly associated with temperature or precipitation across the range and these SNPs were enriched for non-synonymous substitutions. SNPs associated with temperature and aridity were more often found in or near genes than expected by chance and enriched for putative involvement in dormancy processes, seed maturation, response to hyperosmosis and salinity, abscisic acid metabolism, hormone metabolism, and drought recovery.

Conclusions

Both geography and climate adaptation contribute significantly to patterns of genome-wide variation in P.hallii. Population subgroups within P.hallii may represent early stages in the formation of ecotypes. Climate associated loci identified here represent promising targets for future research in this and other perennial grasses.

Crop simulation models are increasingly being used to understand the feasibility of large-scale cellulosic biofuel production along with the multi-dimensional impacts on environmental sustainability. However, how the uncertainty in model parameters impacts model performance for sustainability is unclear. In this case study, sensitivity analyses were conducted for three switchgrass sustainability metrics: total biomass production, nitrogen loss, and soil carbon change using the APEX (Agricultural Policy/Environmental eXtender) model. Fifteen out of the 45 parameters (25 crop growth (CROP) parameters and 20 additional model parameters (PARM)) were identified as influential for the three sustainability metrics for three lowland genotypes (WBC, AP13, and KAN) across two locations (Temple, TX, and Austin, TX). Our sensitivity results showed that parameter importance was not dependent on the genotypes but depended on the variables of interest, and differed only slightly between locations. Influential belowground-related CROP and PARM parameters were identified for each sustainability metric, indicating that belowground-related parameters are just as important as commonly measured aboveground CROP parameters. Further investigation of the linear or non-linear relationships and the two-way interactions between each of the individual influential parameters with the three sustainability metrics reflected the functions and characteristics within the APEX model and the interrelations among different processes. Strong interactions between the most influential parameters for total biomass, nitrogen loss, and soil carbon change also highlighted the importance of accurately setting these parameters. Identification of influential model parameters for switchgrass sustainability may help guide field measurements and provide further understanding of the interrelated processes in the APEX model. Furthermore, future field experiments can be designed to measure these influential parameters and understand the non-linear relationships identified between influential parameters and response variables. More accurate model parameterization will help improve APEX model performance and our understanding of the possible underlying physiological mechanisms.

Environmental stress is a major driver of ecological community dynamics and agricultural productivity. This is especially true for soil water availability, because drought is the greatest abiotic inhibitor of worldwide crop yields. Here, we test the genetic basis of drought responses in the genetic model for C₄ perennial grasses, Panicum hallii, through population genomics, field-scale gene-expression (eQTL) analysis, and comparison of two complete genomes. While gene expression networks are dominated by local cis-regulatory elements, we observe three genomic hotspots of unlinked trans-regulatory loci. These regulatory hubs are four times more drought responsive than the genome-wide average. Additionally, cis- and trans-regulatory networks are more likely to have opposing effects than expected under neutral evolution, supporting a strong influence of compensatory evolution and stabilizing selection. These results implicate trans-regulatory evolution as a driver of drought responses and demonstrate the potential for crop improvement in drought-prone regions through modification of gene regulatory networks.

Biofuels derived from lignocellulosic plant material are an important component of current renewable energy strategies. Improvement eforts in biofuel feedstock crops have been primarily focused on increasing biomass yield with less consideration for tissue quality or composition. Four primary components found in the plant cell wall contribute to the overall quality of plant tissue and conversion characteristics, cellulose and hemicellulose polysaccharides are the primary targets for fuel conversion, while lignin and ash provide structure and defense. We explore the genetic architecture of tissue characteristics using a quantitative trait loci (QTL) mapping approach in Panicum hallii, a model lignocellulosic grass system. Diversity in the mapping population was generated by crossing xeric and mesic varietals, comparative to northern upland and southern lowland ecotypes in switchgrass. We use near-infrared spectroscopy with a primary analytical method to create a P. hallii specifc calibration model to quickly quantify cell wall components.

Immense floral trait variation has likely arisen as an adaptation to attract pollinators. Different pollinator syndromes—suites of floral traits that attract specific pollinator functional groups—are repeatedly observed across closely related taxa or divergent populations. The observation of these trait syndromes suggests that pollinators use floral cues to signal the underlying nectar reward, and that complex trait combinations may persist and evolve through genetic correlations. Here, we explore pollinator preferences and the genetic  architecture of floral divergence using an extensive genetic mapping study in the hybrid zone of two Ipomopsis aggregata subspecies that exhibit a hummingbird and a hawkmoth pollinator syndrome. We found that natural selection acts on several floral traits, and that hummingbirds and hawkmoths exhibited flower color preferences as predicted by their respective pollinator syndromes. Our quantitative trait loci (QTL) analyses revealed 46 loci affecting floral features, many of which co-localize across the genome. Two of these QTL have large effects explaining > 15 percent of the phenotypic variance. The strongest QTL was associated with flower color and localized to a SNP in the anthocyanin biosynthesis pathway (ABP) gene, dihydroflavonol-4-reductase (DFR). Further analysis revealed strong associations between DFR SNP variants, gene  expression and flower color across populations from the hybrid zone. Hence, DFR may be a target of pollinatormediated selection in the hybrid zone of these two subspecies. Together, our findings suggest that hummingbirds and hawkmoths exhibit contrasting flower color preferences, which may drive the divergence of several floral traits through correlated trait evolution.

Reinforcement contact zones, which are secondary contact zones where species are diverging in reproductive behaviors due to selection against hybridization, represent natural laboratories for studying speciation-in-action. Here, we examined replicate localities across the entire reinforcement contact zone between North American chorus frogs Pseudacris feriarum and P. nigrita to investigate geographic variation in hybridization frequencies and to assess whether reinforcement may have contributed to increased genetic divergence within species. Previous work indicated these species have undergone reproductive character displacement (RCD) in male acoustic signals and female preferences due to reinforcement. We also examined acoustic signal variation
across the contact zone to assess whether signal characteristics reliably predict
hybrid index and to elucidate whether the degree of RCD predicts hybridization rate. Using microsatellites, mitochondrial sequences, and acoustic signal information from >1,000 individuals across >50 localities and ten sympatric focal regions, we demonstrate: (1) hybridization occurs and (2) varies substantially across the geographic range of the contact zone, (3) hybridization is asymmetric and in the direction predicted from observed patterns of asymmetric RCD, (4) in one species, genetic distance is higher between conspecific localities where one or both have been reinforced than between nonreinforced localities, after controlling for geographic distance, (5) acoustic signal characters strongly predict hybrid index, and (6) the degree of RCD does not strongly predict admixture levels. By showing that hybridization occurs in all sympatric localities,
this study provides the fifth and final line of evidence that reproductive character
displacement is due to reinforcement in the chorus frog contact zone.  Furthermore, this work suggests that the dual action of cascade reinforcement and partial geographic isolation is promoting genetic diversification within one of the reinforced species.

While prokaryotic pan-genomes have been shown to contain many more genes than any individual organism, the prevalence and functional significance of differentially present genes in eukaryotes remains poorly understood. Whole-genome de novo assembly and annotation of 54 lines of the grass Brachypodium distachyon yield a pan-genome containing nearly twice the number of genes found in any individual genome. Genes present in all lines are enriched for essential biological functions, while genes present in only some lines are enriched for conditionally beneficial functions (e.g., defense and development), display faster evolutionary rates, lie closer to transposable elements and are less likely to be syntenic with orthologous genes in other grasses. Our data suggest that differentially present genes contribute substantially to phenotypic variation within a eukaryote species, these genes have a major influence in population genetics, and transposable elements play a key role in pan-genome evolution.

Global increase in salinity levels has made it imperative to identify novel sources of genetic variation
for tolerance traits, especially in rice. The rice landrace Horkuch, endemic to the saline coastal area of
Bangladesh, was used in this study as the source of tolerance in reciprocal crosses with the sensitive but
high-yielding IR29 variety for discovering transcriptional variation associated with salt tolerance in the
resulting populations. The cytoplasmic effect of the Horkuch background in leaves under stress showed
functional enrichment for signal transduction, DNA-dependent regulation and transport activities. In
roots the enrichment was for cell wall organization and macromolecule biosynthesis. In contrast, the
cytoplasmic effect of IR29 showed upregulation of apoptosis and downregulation of phosphorylation
across tissues relative to Horkuch. Differential gene expression in leaves of the sensitive population
showed downregulation of GO processes like photosynthesis, ATP biosynthesis and ion transport. Roots
of the tolerant plants conversely showed upregulation of GO terms like G-protein coupled receptor
pathway, membrane potential and cation transport. Furthermore, genes involved in regulating
membrane potentials were constitutively expressed only in the roots of tolerant individuals. Overall
our work has developed genetic resources and elucidated the likely mechanisms associated with the
tolerance response of the Horkuch genotype.

Accumulation of the stress hormone abscisic acid (ABA) in response
to drought and low water-potential controls many downstream
acclimation mechanisms. However, mechanisms controlling ABA
accumulation itself are less known. There was a 10-fold range of
variation in ABA levels among nearly 300 Arabidopsis thaliana accessions
exposed to the same low water-potential severity. Genomewide
association analysis (GWAS) identified genomic regions containing
clusters of ABA-associated SNPs. Candidate genes within
these regions included few genes with known stress or ABArelated
function. The GWAS data were used to guide reverse genetic
analysis, which found effectors of ABA accumulation. These included
plasma-membrane–localized signaling proteins such as receptor-like
kinases, aspartic protease, a putative lipid-binding START domain
protein, and other membrane proteins of unknown function as well
as a RING U-box protein and possible effect of tonoplast transport on
ABA accumulation. Putative loss-of-function polymorphisms within
the START domain protein were associated with climate factors at
accession sites of origin, indicating its potential involvement in
drought adaptation. Overall, using ABA accumulation as a basis for
a combined GWAS–reverse genetic strategy revealed the broad natural
variation in low-water-potential–induced ABA accumulation and
was successful in identifying genes that affect ABA levels and may
act in upstream drought-related sensing and signaling mechanisms.
ABA effector loci were identified even when each one was of incremental
effect, consistent with control of ABA accumulation being
distributed among the many branches of ABA metabolism or mediated
by genes with partially redundant function.

An enduring question in plant physiology and evolution is how single genotypes of plants
optimize performance in diverse, often highly variable, environments.
We grew 35 natural accessions of the grass Brachypodium distachyon in four environments
in the glasshouse, contrasting soil water deficit, elevated temperature and their interaction.
We modeled treatment, genotype and interactive effects on leaf-level and whole-plant traits,
including fecundity. We also assessed the relationship between glasshouse-measured traits
and parameters related to climate at the place of origin.
We found abundant genetic variation in both constitutive and induced traits related to
plant–water relations. Most traits showed strong interaction between temperature and water
availability, and we observed genotype-by-environment interaction for several traits. Notably,
leaf free proline abundance showed a strong effect of genotype 9 temperature 9 water. We
found strong associations between phenology, biomass and water use efficiency (WUE) with
parameters describing climate of origin.
Plants respond to multiple stressors in ways not directly predictable from single stressors,
underscoring the complex and trait-specific mechanisms of environmental response. Climate–
trait correlations support a role for WUE and phenology in local adaptation to climate in
B. distachyon.