Reduced free asparagine in wheat grain resulting from a natural deletion of TaASN-B2: investigating and exploiting diversity in the asparagine synthetase gene family to improve wheat quality

Background Understanding the determinants of free asparagine concentration in wheat grain is necessary to reduce levels of the processing contaminant acrylamide in baked and toasted wheat products. Although crop management strategies can help reduce asparagine concentrations, breeders have limited options to select for genetic variation underlying this trait. Asparagine synthetase enzymes catalyse a critical step in asparagine biosynthesis in plants and, in wheat, are encoded by five homeologous gene triads that exhibit distinct expression profiles. Within this family, TaASN2 genes are highly expressed during grain development but TaASN-B2 is absent in some varieties. Results Natural genetic diversity in the asparagine synthetase gene family was assessed in different wheat varieties revealing instances of presence/absence variation and other polymorphisms, including some predicted to affect the function of the encoded protein. The presence and absence of TaASN-B2 was determined across a range of UK and global common wheat varieties and related species, showing that the deletion encompassing this gene was already present in some wild emmer wheat genotypes. Expression profiling confirmed that TaASN2 transcripts were only detectable in the grain, while TaASN3.1 genes were highly expressed during the early stages of grain development. TaASN-A2 was the most highly expressed TaASN2 homeologue in most assayed wheat varieties. TaASN-B2 and TaASN-D2 were expressed at similar, lower levels in varieties possessing TaASN-B2. Expression of TaASN-A2 and TaASN-D2 did not increase to compensate for the absence of TaASN-B2, so total TaASN2 expression was lower in varieties lacking TaASN-B2. Consequently, free asparagine concentrations in field-produced grain were, on average, lower in varieties lacking TaASN-B2, although the effect was lost when free asparagine accumulated to very high concentrations as a result of sulphur deficiency. Conclusions Selecting wheat genotypes lacking the TaASN-B2 gene may be a simple and rapid way for breeders to reduce free asparagine concentrations in commercial wheat grain.

Reduced free asparagine in wheat grain resulting from a natural deletion of TaASN-B2: investigating and exploiting diversity in the asparagine synthetase gene family to improve wheat quality

Background: Understanding the determinants of free asparagine concentration in wheat grain is necessary to reduce levels of the processing contaminant acrylamide in baked and toasted wheat products. Although crop management strategies can help reduce asparagine levels, breeders have limited options to select for genetic variation underlying this trait. Asparagine synthetase enzymes catalyse a critical step in asparagine biosynthesis in plants and, in wheat, are encoded by five homeologous gene triads that exhibit distinct expression profiles. Within this family, TaASN2 genes are highly expressed during grain development but TaASN-B2 is absent in some varieties. Results: Natural genetic diversity in the asparagine synthetase gene family was assessed in different wheat varieties revealing instances of presence/absence variation and other polymorphisms, including some predicted to affect the function of the encoded protein. The presence and absence of TaASN-B2 was determined across a range of UK and global common wheat varieties and related species, showing that the deletion encompassing this gene was already present in some wild emmer wheat genotypes. Expression profiling confirmed that TaASN2 transcripts were only detectable in the grain, while TaASN3.1 genes were highly expressed during the early stages of grain development. TaASN-A2 was the most highly expressed TaASN2 homeologue in most assayed wheat varieties. TaASN-B2 and TaASN-D2 were expressed at similar, lower levels in varieties possessing TaASN-B2. Expression of TaASN-A2 and TaASN-D2 did not increase to compensate for the absence of TaASN-B2, so total TaASN2 expression was lower in varieties lacking TaASN-B2. Consequently, free asparagine levels in field-produced grain were, on average, lower in varieties lacking TaASN-B2, although the effect was lost when free asparagine accumulated to very high levels as a result of sulphur deficiency.Conclusions: Selecting wheat genotypes lacking the TaASN-B2 gene may be a simple and rapid way for breeders to reduce free asparagine levels in commercial wheat grain.

Evolution of Homeologous Gene Expression in Polyploid Wheat

Polyploidization has played a prominent role in the evolutionary history of plants. Two recent and sequential allopolyploidization events have resulted in the formation of wheat species with different ploidies, and which provide a model to study the effects of polyploidization on the evolution of gene expression. In this study, we identified differentially expressed genes (DEGs) between four BBAA tetraploid wheats of three different ploidy backgrounds. DEGs were found to be unevenly distributed among functional categories and duplication modes. We observed more DEGs in the extracted tetraploid wheat (ETW) than in natural tetraploid wheats (TD and TTR13) as compared to a synthetic tetraploid (AT2). Furthermore, DEGs showed higher Ka/Ks ratios than those that did not show expression changes (non-DEGs) between genotypes, indicating DEGs and non-DEGs experienced different selection pressures. For A-B homeolog pairs with DEGs, most of them had only one differentially expressed copy, however, when both copies of a homeolog pair were DEGs, the A and B copies were more likely to be regulated to the same direction. Our results suggest that both cis- and inter-subgenome trans-regulatory changes are important drivers in the evolution of homeologous gene expression in polyploid wheat, with ploidy playing a significant role in the process.

A subfunctionalization epistasis model to evaluate homeologous gene interactions in allopolyploid wheat

1AbstractHybridization between related species results in the formation of an allopolyploid with multiple subgenomes. These subgenomes will each contain complete, yet evolutionarily divergent, sets of genes. Like a diploid hybrid, allopolyploids will have two versions, or homeoalleles, for every gene. Partial functional redundancy between homeologous genes should result in a deviation from additivity. These epistatic interactions between homeoalleles are analogous to dominance effects, but are fixed across subgenomes through self pollination. An allopolyploid can be viewed as an immortalized hybrid, with the opportunity to select and fix favorable homeoallelic interactions within inbred varieties. We present a subfunctionalization epistasis model to estimate the degree of functional redundancy between homeoallelic loci and a statistical framework to determine their importance within a population. We provide an example using the homeologous dwarfing genes of allohexaploid wheat, Rht-1, and search for genome-wide patterns indicative of homeoallelic subfunctionalization in a breeding population. Using the IWGSC RefSeq vl.0 sequence, 23,796 homeoallelic gene sets were identified and anchored to the nearest DNA marker to form 10,172 homeologous marker sets. Interaction predictors constructed from products of marker scores were used to fit the homeologous main and interaction effects, as well as estimate whole genome genetic values. Some traits displayed a pattern indicative of homeoallelic subfunctionalization, while other traits showed a less clear pattern or were not affected. Using genomic prediction accuracy to evaluate importance of marker interactions, we show that homeologous interactions explain a portion of the non-additive genetic signal, but are less important than other epistatic interactions.

Mismatch correction acts as a barrier to homeologous recombination in Saccharomyces cerevisiae.

A homeologous mitotic recombination assay was used to test the role of Saccharomyces cerevisiae mismatch repair genes PMS1, MSH2 and MSH3 on recombination fidelity. A homeologous gene pair consisting of S. cerevisiae SPT15 and its S. pombe homolog were present as a direct repeat on chromosome V, with the exogenous S. pombe sequences inserted either upstream or downstream of the endogenous S. cerevisiae gene. Each gene carried a different inactivating mutation, rendering the starting strain Spt15-. Recombinants that regenerated SPT15 function were scored after nonselective growth of the cells. In strains wild type for mismatch repair, homeologous recombination was depressed 150- to 180-fold relative to homologous controls, indicating that recombination between diverged sequences is inhibited. In one orientation of the homeologous gene pair, msh2 or msh3 mutations resulted in 17- and 9.6-fold elevations in recombination and the msh2 msh3 double mutant exhibited an 43-fold increase, implying that each MSH gene can function independently in trans to prevent homeologous recombination. Homologous recombination was not significantly affected by the msh mutations. In the other orientation, only msh2 strains were elevated (12-fold) for homeologous recombination. A mutation in MSH3 did not affect the rate of recombination in this orientation. Surprisingly, a pms1 deletion mutant did not exhibit elevated homeologous recombination.