Evidence that variation in root anatomy contributes to local adaptation in Mexican native maize.

Mexican native maize (Zea mays ssp. mays) is adapted to a wide range of climatic and edaphic conditions. Here, we focus specifically on the potential role of root anatomical variation in this adaptation. Given the investment required to characterize root anatomy, we present a machine-learning approach using environmental descriptors to project trait variation from a relatively small training panel onto a larger panel of genotyped and georeferenced Mexican maize accessions. The resulting models defined potential biologically relevant clines across a complex environment that we used subsequently for genotype-environment association. We found evidence of systematic variation in maize root anatomy across Mexico, notably a prevalence of trait combinations favoring a reduction in axial hydraulic conductance in varieties sourced from cooler, drier highland areas. We discuss our results in the context of previously described water-banking strategies and present candidate genes that are associated with both root anatomical and environmental variation. Our strategy is a refinement of standard environmental genome-wide association analysis that is applicable whenever a training set of georeferenced phenotypic data is available.

Genetic analysis and QTL mapping of domestication-related traits in chili pepper (Capsicum annuum L.).

Chili pepper (Capsicum annuum L.) is one of the oldest and most phenotypically diverse pre-Columbian crops of the Americas. Despite the abundance of genetic resources, the use of wild germplasm and landraces in chili pepper breeding is limited. A better understanding of the evolutionary history in chili peppers, particularly in the context of traits of agronomic interest, can contribute to future improvement and conservation of genetic resources. In this study, an F2:3 mapping population derived from a cross between a C. annuum wild accession (Chiltepin) and a cultivated variety (Puya) was used to identify genomic regions associated with 19 domestication and agronomic traits. A genetic map was constructed consisting of 1023 single nucleotide polymorphism (SNP) markers clustered into 12 linkage groups and spanning a total of 1,263.87 cM. A reciprocal translocation that differentiates the domesticated genome from its wild ancestor and other related species was identified between chromosomes 1 and 8. Quantitative trait locus (QTL) analysis detected 20 marker-trait associations for 13 phenotypes, from which 14 corresponded to previously identified loci, and six were novel genomic regions related to previously unexplored domestication-syndrome traits, including form of unripe fruit, seedlessness, deciduous fruit, and growth habit. Our results revealed that the genetic architecture of Capsicum domestication is similar to other domesticated species with few loci with large effects, the presence of QTLs clusters in different genomic regions, and the predominance of domesticated recessive alleles. Our analysis indicates the domestication process in chili pepper has also had an effect on traits not directly related to the domestication syndrome. The information obtained in this study provides a more complete understanding of the genetic basis of Capsicum domestication that can potentially guide strategies for the exploitation of wild alleles.

Genetic mapping of maize metabolites using high-throughput mass profiling.

Plant metabolites are the basis of human nutrition and have biological relevance in ecology. Farmers selected plants with favorable characteristics since prehistoric times and improved the cultivars, but without knowledge of underlying mechanisms. Understanding the genetic basis of metabolite production can facilitate the successful breeding of plants with augmented nutritional value. To identify genetic factors related to the metabolic composition in maize, we generated mass profiles of 198 recombinant inbred lines (RILs) and their parents (B73 and Mo17) using direct-injection electrospray ionization mass spectrometry (DLI-ESI MS). Mass profiling allowed the correct clustering of samples according to genotype. We quantified 71 mass features from grains and 236 mass features from leaf extracts. For the corresponding ions, we identified tissue-specific metabolic 'Quantitative Trait Loci' (mQTLs) distributed across the maize genome. These genetic regions could regulate multiple metabolite biosynthesis pathways. Our findings demonstrate that DLI-ESI MS has sufficient analytical resolution to map mQTLs. These identified genetic loci will be helpful in metabolite-focused maize breeding. Mass profiling is a powerful tool for detecting mQTLs in maize and enables the high-throughput screening of loci responsible for metabolite biosynthesis.

An adaptive teosinte mexicana introgression modulates phosphatidylcholine levels and is associated with maize flowering time.

Native Americans domesticated maize (Zea mays ssp. mays) from lowland teosinte parviglumis (Zea mays ssp. parviglumis) in the warm Mexican southwest and brought it to the highlands of Mexico and South America where it was exposed to lower temperatures that imposed strong selection on flowering time. Phospholipids are important metabolites in plant responses to low-temperature and phosphorus availability and have been suggested to influence flowering time. Here, we combined linkage mapping with genome scans to identify High PhosphatidylCholine 1 (HPC1), a gene that encodes a phospholipase A1 enzyme, as a major driver of phospholipid variation in highland maize. Common garden experiments demonstrated strong genotype-by-environment interactions associated with variation at HPC1, with the highland HPC1 allele leading to higher fitness in highlands, possibly by hastening flowering. The highland maize HPC1 variant resulted in impaired function of the encoded protein due to a polymorphism in a highly conserved sequence. A meta-analysis across HPC1 orthologs indicated a strong association between the identity of the amino acid at this position and optimal growth in prokaryotes. Mutagenesis of HPC1 via genome editing validated its role in regulating phospholipid metabolism. Finally, we showed that the highland HPC1 allele entered cultivated maize by introgression from the wild highland teosinte Zea mays ssp. mexicana and has been maintained in maize breeding lines from the Northern United States, Canada, and Europe. Thus, HPC1 introgressed from teosinte mexicana underlies a large metabolic QTL that modulates phosphatidylcholine levels and has an adaptive effect at least in part via induction of early flowering time.

A B73×Palomero Toluqueño mapping population reveals local adaptation in Mexican highland maize.

Generations of farmer selection in the central Mexican highlands have produced unique maize varieties adapted to the challenges of the local environment. In addition to possessing great agronomic and cultural value, Mexican highland maize represents a good system for the study of local adaptation and acquisition of adaptive phenotypes under cultivation. In this study, we characterize a recombinant inbred line population derived from the B73 reference line and the Mexican highland maize variety Palomero Toluqueño. B73 and Palomero Toluqueño showed classic rank-changing differences in performance between lowland and highland field sites, indicative of local adaptation. Quantitative trait mapping identified genomic regions linked to effects on yield components that were conditionally expressed depending on the environment. For the principal genomic regions associated with ear weight and total kernel number, the Palomero Toluqueño allele conferred an advantage specifically in the highland site, consistent with local adaptation. We identified Palomero Toluqueño alleles associated with expression of characteristic highland traits, including reduced tassel branching, increased sheath pigmentation and the presence of sheath macrohairs. The oligogenic architecture of these three morphological traits supports their role in adaptation, suggesting they have arisen from consistent directional selection acting at distinct points across the genome. We discuss these results in the context of the origin of phenotypic novelty during selection, commenting on the role of de novo mutation and the acquisition of adaptive variation by gene flow from endemic wild relatives.

Low nitrogen availability inhibits the phosphorus starvation response in maize (Zea mays ssp. mays L.).

BACKGROUND: Nitrogen (N) and phosphorus (P) are macronutrients essential for crop growth and productivity. In cultivated fields, N and P levels are rarely sufficient, contributing to the gap between realized and potential production. Fertilizer application increases nutrient availability, but is not available to all farmers, nor are current rates of application sustainable or environmentally desirable. Transcriptomic studies of cereal crops have revealed dramatic responses to either low N or low P single stress treatments. In the field, however, levels of both N and P may be suboptimal. The interaction between N and P starvation responses remains to be fully characterized. RESULTS: We characterized growth and root and leaf transcriptomes of young maize plants under nutrient replete, low N, low P or combined low NP conditions. We identified 1555 genes to respond to our nutrient treatments, in one or both tissues. A large group of genes, including many classical P starvation response genes, were regulated antagonistically between low N and P conditions. An additional experiment over a range of N availability indicated that a mild reduction in N levels was sufficient to repress the low P induction of P starvation genes. Although expression of P transporter genes was repressed under low N or low NP, we confirmed earlier reports of P hyper accumulation under N limitation. CONCLUSIONS: Transcriptional responses to low N or P were distinct, with few genes responding in a similar way to the two single stress treatments. In combined NP stress, the low N response dominated, and the P starvation response was largely suppressed. A mild reduction in N availability was sufficient to repress the induction of P starvation associated genes. We conclude that activation of the transcriptional response to P starvation in maize is contingent on N availability.

Identification of the maize Mediator CDK8 module and transposon-mediated mutagenesis of ZmMed12a.

Mediator is a conserved transcriptional co-activator that links transcription factors bound at enhancer elements to RNA Polymerase II. Mediator-RNA Polymerase II interactions can be sterically hindered by the Cyclin Dependent Kinase 8 (CDK8) module, a submodule of Mediator that acts to repress transcription in response to discrete cellular and environmental cues. The CDK8 module is conserved in all eukaryotes and consists of 4 proteins: CDK8, CYCLIN C (CYCC), MED12, and MED13. In this study, we have characterized the CDK8 module of Mediator in maize using genomic, molecular and functional resources. The maize genome contains single copy genes for Cdk8, CycC, and Med13, and two genes for Med12. Analysis of expression data for the CDK8 module demonstrated that all five genes are broadly expressed in maize tissues, and change their expression in response to phosphate and nitrogen limitation. We performed Dissociation (Ds) insertional mutagenesis, recovering two independent insertions in the ZmMed12a gene, one of which produces a truncated transcript. Our molecular identification of the maize CDK8 module, assays of CDK8 module expression under nutrient limitation, and characterization of transposon insertions in ZmMed12a establish the basis for molecular and functional studies of the role of these important transcriptional regulators in development and nutrient homeostasis in Zea mays.

The Phosphoglycerate Kinase (PGK) Gene Family of Maize (Zea mays var. B73).

Phosphoglycerate kinase (PGK, E.C. 2.7.2.3) interconverts ADP + 1,3-bisphospho-glycerate (1,3-bPGA) to ATP + 3-phosphoglycerate (3PGA). While most bacteria have a single pgk gene and mammals possess two copies, plant genomes contain three or more PGK genes. In this study, we identified five Pgk genes in the Zea mays var. B73 genome, predicted to encode proteins targeted to different subcellular compartments: ZmPgk1, ZmPgk2, and ZmPgk4 (chloroplast), ZmPgk3 (cytosol), and ZmPgk5 (nucleus). The expression of ZmPgk3 was highest in non-photosynthetic tissues (roots and cobs), where PGK activity was also greatest, consistent with a function in glycolysis. Green tissues (leaf blade and husk leaf) showed intermediate levels of PGK activity, and predominantly expressed ZmPgk1 and ZmPgk2, suggesting involvement in photosynthetic metabolism. ZmPgk5 was weakly expressed and ZmPgk4 was not detected in any tissue. Phylogenetic analysis showed that the photosynthetic and glycolytic isozymes of plants clustered together, but were distinct from PGKs of animals, fungi, protozoa, and bacteria, indicating that photosynthetic and glycolytic isozymes of plants diversified after the divergence of the plant lineage from other groups. These results show the distinct role of each PGK in maize and provide the basis for future studies into the regulation and function of this key enzyme.

The genetic architecture of host response reveals the importance of arbuscular mycorrhizae to maize cultivation.

Arbuscular mycorrhizal fungi (AMF) are ubiquitous in cultivated soils, forming symbiotic relationships with the roots of major crop species. Studies in controlled conditions have demonstrated the potential of AMF to enhance the growth of host plants. However, it is difficult to estimate the actual benefit in the field, not least because of the lack of suitable AMF-free controls. Here we implement a novel strategy using the selective incorporation of AMF-resistance into a genetic mapping population to evaluate maize response to AMF. We found AMF to account for about one-third of the grain production in a medium input field, as well as to affect the relative performance of different plant genotypes. Characterization of the genetic architecture of the host response indicated a trade-off between mycorrhizal dependence and benefit. We identified several QTL linked to host benefit, supporting the feasibility of breeding crops to maximize profit from symbiosis with AMF.

Characterization of introgression from the teosinte Zea mays ssp. mexicana to Mexican highland maize.

BACKGROUND: The spread of maize cultivation to the highlands of central Mexico was accompanied by substantial introgression from the endemic wild teosinte Zea mays ssp. mexicana, prompting the hypothesis that the transfer of beneficial variation facilitated local adaptation. METHODS: We used whole-genome sequence data to map regions of Zea mays ssp. mexicana introgression in three Mexican highland maize individuals. We generated a genetic linkage map and performed Quantitative Trait Locus mapping in an F2 population derived from a cross between lowland and highland maize individuals. RESULTS: Introgression regions ranged in size from several hundred base pairs to Megabase-scale events. Gene density within introgression regions was comparable to the genome as a whole, and over 1,000 annotated genes were located within introgression events. Quantitative Trait Locus mapping identified a small number of loci linked to traits characteristic of Mexican highland maize. DISCUSSION: Although there was no strong evidence to associate quantitative trait loci with regions of introgression, we nonetheless identified many Mexican highland alleles of introgressed origin that carry potentially functional sequence variants. The impact of introgression on stress tolerance and yield in the highland environment remains to be fully characterized.

Allele specific expression analysis identifies regulatory variation associated with stress-related genes in the Mexican highland maize landrace Palomero Toluqueño.

BACKGROUND: Gene regulatory variation has been proposed to play an important role in the adaptation of plants to environmental stress. In the central highlands of Mexico, farmer selection has generated a unique group of maize landraces adapted to the challenges of the highland niche. In this study, gene expression in Mexican highland maize and a reference maize breeding line were compared to identify evidence of regulatory variation in stress-related genes. It was hypothesised that local adaptation in Mexican highland maize would be associated with a transcriptional signature observable even under benign conditions. METHODS: Allele specific expression analysis was performed using the seedling-leaf transcriptome of an F1 individual generated from the cross between the highland adapted Mexican landrace Palomero Toluqueño and the reference line B73, grown under benign conditions. Results were compared with a published dataset describing the transcriptional response of B73 seedlings to cold, heat, salt and UV treatments. RESULTS: A total of 2,386 genes were identified to show allele specific expression. Of these, 277 showed an expression difference between Palomero Toluqueño and B73 alleles under benign conditions that anticipated the response of B73 cold, heat, salt and/or UV treatments, and, as such, were considered to display a prior stress response. Prior stress response candidates included genes associated with plant hormone signaling and a number of transcription factors. Construction of a gene co-expression network revealed further signaling and stress-related genes to be among the potential targets of the transcription factors candidates. DISCUSSION: Prior activation of responses may represent the best strategy when stresses are severe but predictable. Expression differences observed here between Palomero Toluqueño and B73 alleles indicate the presence of cis-acting regulatory variation linked to stress-related genes in Palomero Toluqueño. Considered alongside gene annotation and population data, allele specific expression analysis of plants grown under benign conditions provides an attractive strategy to identify functional variation potentially linked to local adaptation.

Phosphorus acquisition efficiency in arbuscular mycorrhizal maize is correlated with the abundance of root-external hyphae and the accumulation of transcripts encoding PHT1 phosphate transporters.

Plant interactions with arbuscular mycorrhizal fungi have long attracted interest for their potential to promote more efficient use of mineral resources in agriculture. Their use, however, remains limited by a lack of understanding of the processes that determine the outcome of the symbiosis. In this study, the impact of host genotype on growth response to mycorrhizal inoculation was investigated in a panel of diverse maize lines. A panel of 30 maize lines was evaluated with and without inoculation with arbuscular mycorrhizal fungi. The line Oh43 was identified to show superior response and, along with five other reference lines, was characterized in greater detail in a split-compartment system, using 33 P to quantify mycorrhizal phosphorus uptake. Changes in relative growth indicated variation in host capacity to profit from the symbiosis. Shoot phosphate content, abundance of root-internal and -external fungal structures, mycorrhizal phosphorus uptake, and accumulation of transcripts encoding plant PHT1 family phosphate transporters varied among lines. Superior response in Oh43 is correlated with extensive development of root-external hyphae, accumulation of specific Pht1 transcripts and high phosphorus uptake by mycorrhizal plants. The data indicate that host genetic factors influence fungal growth strategy with an impact on plant performance.

The maize (Zea mays ssp. mays var. B73) genome encodes 33 members of the purple acid phosphatase family.

Purple acid phosphatases (PAPs) play an important role in plant phosphorus nutrition, both by liberating phosphorus from organic sources in the soil and by modulating distribution within the plant throughout growth and development. Furthermore, members of the PAP protein family have been implicated in a broader role in plant mineral homeostasis, stress responses and development. We have identified 33 candidate PAP encoding gene models in the maize (Zea mays ssp. mays var. B73) reference genome. The maize Pap family includes a clear single-copy ortholog of the Arabidopsis gene AtPAP26, shown previously to encode both major intracellular and secreted acid phosphatase activities. Certain groups of PAPs present in Arabidopsis, however, are absent in maize, while the maize family contains a number of expansions, including a distinct radiation not present in Arabidopsis. Analysis of RNA-sequencing based transcriptome data revealed accumulation of maize Pap transcripts in multiple plant tissues at multiple stages of development, and increased accumulation of specific transcripts under low phosphorus availability. These data suggest the maize PAP family as a whole to have broad significance throughout the plant life cycle, while highlighting potential functional specialization of individual family members.