Targeted Mutagenesis of the Multicopy Myrosinase Gene Family in Allotetraploid Brassica juncea Reduces Pungency in Fresh Leaves across Environments.

Recent breeding efforts in Brassica have focused on the development of new oilseed feedstock crop for biofuels (e.g., ethanol, biodiesel, bio-jet fuel), bio-industrial uses (e.g., bio-plastics, lubricants), specialty fatty acids (e.g., erucic acid), and producing low glucosinolates levels for oilseed and feed meal production for animal consumption. We identified a novel opportunity to enhance the availability of nutritious, fresh leafy greens for human consumption. Here, we demonstrated the efficacy of disarming the 'mustard bomb' reaction in reducing pungency upon the mastication of fresh tissue-a major source of unpleasant flavor and/or odor in leafy Brassica. Using gene-specific mutagenesis via CRISPR-Cas12a, we created knockouts of all functional copies of the type-I myrosinase multigene family in tetraploid Brassica juncea. Our greenhouse and field trials demonstrate, via sensory and biochemical analyses, a stable reduction in pungency in edited plants across multiple environments. Collectively, these efforts provide a compelling path toward boosting the human consumption of nutrient-dense, fresh, leafy green vegetables.

Ecological factors influence balancing selection on leaf chemical profiles of a wildflower.

Balancing selection is frequently invoked as a mechanism that maintains variation within and across populations. However, there are few examples of balancing selection operating on loci underpinning complex traits, which frequently display high levels of variation. We investigated mechanisms that may maintain variation in a focal polymorphism-leaf chemical profiles of a perennial wildflower (Boechera stricta, Brassicaceae)-explicitly interrogating multiple ecological and genetic processes including spatial variation in selection, antagonistic pleiotropy and frequency-dependent selection. A suite of common garden and greenhouse experiments showed that the alleles underlying variation in chemical profile have contrasting fitness effects across environments, implicating two ecological drivers of selection on chemical profile: herbivory and drought. Phenotype-environment associations and molecular genetic analyses revealed additional evidence of past selection by these drivers. Together, these data are consistent with balancing selection on chemical profile, probably caused by pleiotropic effects of secondary chemical biosynthesis genes on herbivore defence and drought response.

Correction to: Ancient polymorphisms contribute to genome-wide variation by long-term balancing selection and divergent sorting in Boechera stricta.

Following publication of the original article [1], the authors reported that the Availability of data and materials section required updating. The updated text reads as follows.

Ancient polymorphisms contribute to genome-wide variation by long-term balancing selection and divergent sorting in Boechera stricta.

BACKGROUND: Genomic variation is widespread, and both neutral and selective processes can generate similar patterns in the genome. These processes are not mutually exclusive, so it is difficult to infer the evolutionary mechanisms that govern population and species divergence. Boechera stricta is a perennial relative of Arabidopsis thaliana native to largely undisturbed habitats with two geographic and ecologically divergent subspecies. Here, we delineate the evolutionary processes driving the genetic diversity and population differentiation in this species. RESULTS: Using whole-genome re-sequencing data from 517 B. stricta accessions, we identify four genetic groups that diverged around 30-180 thousand years ago, with long-term small effective population sizes and recent population expansion after the Last Glacial Maximum. We find three genomic regions with elevated nucleotide diversity, totaling about 10% of the genome. These three regions of elevated nucleotide diversity show excess of intermediate-frequency alleles, higher absolute divergence (dXY), and lower relative divergence (FST) than genomic background, and significant enrichment in immune-related genes, reflecting long-term balancing selection. Scattered across the genome, we also find regions with both high FST and dXY among the groups, termed FST-islands. Population genetic signatures indicate that FST-islands with elevated divergence, which have experienced directional selection, are derived from divergent sorting of ancient polymorphisms. CONCLUSIONS: Our results suggest that long-term balancing selection on disease resistance genes may have maintained ancestral haplotypes across different geographical lineages, and unequal sorting of balanced polymorphisms may have generated genomic regions with elevated divergence. This study highlights the importance of ancestral balanced polymorphisms as crucial components of genome-wide variation.

Publisher correction: Young inversion with multiple linked QTLs under selection in a hybrid zone.

In Fig. 5 of the version of this Article originally published, the final number on the x axes of each panel was incorrectly written as 1.5; it should have read 7.5. This has now been corrected in all versions of the Article.

Young inversion with multiple linked QTLs under selection in a hybrid zone.

Fixed chromosomal inversions can reduce gene flow and promote speciation in two ways: by suppressing recombination and by carrying locally favoured alleles at multiple loci. However, it is unknown whether favoured mutations slowly accumulate on older inversions or if young inversions spread because they capture pre-existing adaptive quantitative trait loci (QTLs). By genetic mapping, chromosome painting and genome sequencing, we have identified a major inversion controlling ecologically important traits in Boechera stricta. The inversion arose since the last glaciation and subsequently reached local high frequency in a hybrid speciation zone. Furthermore, the inversion shows signs of positive directional selection. To test whether the inversion could have captured existing, linked QTLs, we crossed standard, collinear haplotypes from the hybrid zone and found multiple linked phenology QTLs within the inversion region. These findings provide the first direct evidence that linked, locally adapted QTLs may be captured by young inversions during incipient speciation.

Field-based high throughput phenotyping rapidly identifies genomic regions controlling yield components in rice.

To ensure food security in the face of population growth, decreasing water and land for agriculture, and increasing climate variability, crop yields must increase faster than the current rates. Increased yields will require implementing novel approaches in genetic discovery and breeding. Here we demonstrate the potential of field-based high throughput phenotyping (HTP) on a large recombinant population of rice to identify genetic variation underlying important traits. We find that detecting quantitative trait loci (QTL) with HTP phenotyping is as accurate and effective as traditional labor-intensive measures of flowering time, height, biomass, grain yield, and harvest index. Genetic mapping in this population, derived from a cross of an modern cultivar (IR64) with a landrace (Aswina), identified four alleles with negative effect on grain yield that are fixed in IR64, demonstrating the potential for HTP of large populations as a strategy for the second green revolution.

Genetics of water use physiology in locally adapted Arabidopsis thaliana.

Identifying the genetic basis of adaptation to climate has long been a goal in evolutionary biology and has applications in agriculture. Adaptation to drought represents one important aspect of local adaptation, and drought is the major factor limiting agricultural yield. We examined local adaptation between Sweden and Italy Arabidopsis thaliana ecotypes, which show contrasting levels of water availability in their local environments. To identify quantitative trait loci (QTL) controlling water use physiology traits and adaptive trait QTL (genomic regions where trait QTL and fitness QTL colocalize), we performed QTL mapping on 374F9 recombinant inbred lines in well-watered and terminal drought conditions. We found 72 QTL (32 in well-watered, 31 in drought, 9 for plasticity) across five water use physiology traits: δ(13)C, rosette area, dry rosette weight, leaf water content and percent leaf nitrogen. Some of these genomic regions colocalize with fitness QTL and with other physiology QTL in defined hotspots. In addition, we found evidence of both constitutive and inducible water use physiology QTL. Finally, we identified highly divergent candidate genes, in silico. Our results suggest that many genes with minor effects may influence adaptation through water use physiology and that pleiotropic water use physiology QTL have fitness consequences.

The genomic signal of partial sweeps in Mimulus guttatus.

The molecular signature of selection depends strongly on whether new mutations are immediately favorable and sweep to fixation (hard sweeps) as opposed to when selection acts on segregating variation (soft sweeps). The prediction of reduced sequence variation around selected polymorphisms is much stronger for hard than soft sweeps, particularly when considering quantitative traits where sweeps are likely to be incomplete. Here, we directly investigate the genomic signal of soft sweeps within an artificial selection experiment on Mimulus guttatus. We first develop a statistical method based on Fisher's angular transformation of allele frequencies to identify selected loci. Application of this method identifies about 400 significant windows, but no fixed differences between phenotypically divergent populations. With two notable exceptions, we find a modest average effect of partial sweeps on the amount of molecular variation. The first exception is a polymorphic inversion on chromosome 6. The increase of the derived haplotype has a broad genomic effect due to recombination suppression coupled with substantial initial haplotype structure within the population. Second, we found significant increases in nucleotide variation around selected loci in the population evolving larger flowers. This suggests that "high" alleles for flower size were initially less frequent than "low" alleles. This result is consistent with prior studies of M. guttatus and illustrates how molecular evolution can depend on the allele frequency spectrum at quantitative trait loci.

Spatially and temporally varying selection on intrapopulation quantitative trait loci for a life history trade-off in Mimulus guttatus.

Why do populations remain genetically variable despite strong continuous natural selection? Mutation reconstitutes variation eliminated by selection and genetic drift, but theoretical and experimental studies each suggest that mutation-selection balance insufficient to explain extant genetic variation in most complex traits. The alternative hypothesis of balancing selection, wherein selection maintains genetic variation, is an aggregate of multiple mechanisms (spatial and temporal heterogeneity in selection, frequency-dependent selection, antagonistic pleiotropy, etc.). Most of these mechanisms have been demonstrated for Mendelian traits, but there is little comparable data for loci affecting quantitative characters. Here, we report a 3-year field study of selection on intrapopulation quantitative trait loci (QTL) of flower size, a highly polygenic trait in Mimulus guttatus. The QTL exhibit antagonistic pleiotropy: alleles that increase flower size, reduce viability, but increase fecundity. The magnitude and direction of selection fluctuates yearly and on a spatial scale of metres. This study provides direct evidence of balancing selection mechanisms on QTL of an ecologically relevant trait.

Interactions among flower-size QTL of Mimulus guttatus are abundant but highly variable in nature.

The frequency and character of interactions among genes influencing complex traits remain unknown. Our ignorance is most acute for segregating variation within natural populations, the epistasis most relevant for quantitative trait evolution. Here, we report a comprehensive survey of interactions among a defined set of flower-size QTL: loci polymorphic within a single natural population of yellow monkeyflower (Mimulus guttatus). We find that epistasis is typical. Observed phenotypes routinely differ from those predicted on the basis of direct allelic affects in the isogenic background, although the direction of deviations is highly variable. Across QTL pairs, there are significantly positive and negative interactions for every trait. Across traits, specific locus pairs routinely exhibit both positive and negative interactions. There was a tendency for negative epistasis to accompany positive direct effects and vice versa for the trait of corolla width, which may be due, at least in part, to the fact that QTL were identified from their direct effects on this trait.

Viability selection prior to trait expression is an essential component of natural selection.

Natural selection operates throughout the life cycle of an organism. Correlative studies typically fail to consider the effects of viability selection prior to trait expression. A 3-year field experiment on the wildflower Mimulus guttatus demonstrates that this unmeasured component of selection can be very strong. As in previous studies, we find that fecundity is positively related to flower size. However, survival to flowering is much lower in large-flowered genotypes than in small-flowered genotypes. Aggregating viability and fecundity, lifetime fitness through female function generally favoured smaller flowered genotypes. This result differs from the great majority of field studies, which suggest strong positive selection on flower size. It has important cautionary implications for studies of natural and sexual selection on adult characters generally, in both plants and animals.