Whole genome sequencing uncovers the structural and transcriptomic landscape of hexaploid wheat/Am. muticum introgression lines

Wheat is a globally vital crop, but its limited genetic variation creates a challenge for breeders aiming to maintain or accelerate agricultural improvements over time. Introducing novel genes and alleles from wheat's wild relatives into the wheat breeding pool via introgression lines is an important component of overcoming this low variation but is limited by poor genomic resolution and limited understanding of the genomic impact of introgression breeding. By sequencing 17 hexaploid wheat/Ambylopyrum muticum introgression lines and the parent lines, we have precisely pinpointed the borders of introgressed segments. We report a genome assembly and annotation of Am. muticum that has facilitated the identification of Am. muticum resistance genes commonly introgressed in lines resistant to stripe rust. Our analysis has identified an abundance of structural disruption and homoeologous pairing across the introgression lines, likely caused by the suppressed Ph1 locus. mRNAseq analysis of six of these introgression lines revealed that introgressed genes tend to be downregulated, shifting the expression balance of triads towards suppression of the introgressed region, with no discernible compensation in the expression of the homoeologous copies. This analysis explores the genomic impact of introgression breeding and provides an affordable way for breeders to better characterise introgression lines and more effectively deploy wild relative variation.

Analysis of Chromosome Associations during Early Meiosis in Wheat Lines Carrying Chromosome Introgressions from Agropyron cristatum

Crested wheatgrass (Agropyron cristatum L. Gaertn., genome P), included in the Triticeae tribe (family Poaceae), is one of the most important grasses in temperate regions. It has been valued as a donor of important agronomic traits for wheat improvement, including tolerance to cold, drought, and high salinity, as well as resistance to leaf rust, stripe rust, and powdery mildew. For successful incorporation of beneficial alleles into wheat, it is essential that recombination between wheat and A. cristatum chromosomes occurs. In this work, we analysed chromosome associations during meiosis in wheat lines carrying chromosome introgressions from A. cristatum chromosomes 5P and 6P in the presence and absence of Ph1 locus using fluorescence in situ hybridisation. The results showed that the Ph1 locus does not affect chromosome associations between A. cristatum and wheat chromosomes because there were no interspecific chromosome associations; therefore, no recombination between chromosomes from wheat and Agropyron were observed in the absence of the Ph1 locus. The 5P and 6P A. cristatum chromosomes do not have a suppressor effect on the Ph1 locus. Wheat univalents in metaphase I suggest that Agropyron chromosomes might carry genes having a role in wheat homologous chromosome associations. Putative effect of the Agropyron genes on wheat chromosome associations does not interact with the Ph1 locus.

Wild and Cultivated Homoeologous Barley Chromosomes Can Associate and Recombine in Wheat in the Absence of the Ph1 Locus

Bread wheat is an allohexaploid that behaves as a diploid during meiosis, the cell division process to produce the gametes occurring in organisms with sexual reproduction. Knowledge of the mechanisms implicated in meiosis can contribute to facilitating the transfer of desirable traits from related species into a crop like wheat in the framework of breeding. It is particularly interesting to shed light on the mechanisms controlling correct pairing between homologous (equivalent) chromosomes and recombination, even more in polyploid species. The Ph1 (Pairing homoeologous 1) locus is implicated in recombination. In this work, we aimed to study whether homoeologous (equivalent chromosomes from different genomes) Hordeum chilense (wild barley) and H. vulgare (cultivated barley) chromosomes can associate and recombine during meiosis in the wheat background in the absence of the Ph1 locus. For this, we have developed H. chilense and H. vulgare double monosomic addition lines for the same and for different homoeology group in wheat in the ph1b mutant background. Using genomic in situ hybridization, we visualized the two (wild and cultivated) barley chromosomes during meiosis and we studied the processes of recognition, association, and recombination between homoeologous chromosomes in the absence of the Ph1 locus. Our results showed that the Ph1 locus does not prevent homoeologous chromosome pairing but it can regulate recombination.

Genome-wide transcription during early wheat meiosis is independent of synapsis, ploidy level and the Ph1 locus

ABSTRACTPolyploidization is a fundamental process in plant evolution. One of the biggest challenges faced by a new polyploid is meiosis, particularly discriminating between multiple related chromosomes so that only homologous chromosomes synapse and recombine to ensure regular chromosome segregation and balanced gametes. Despite its large genome size, high DNA repetitive content and similarity between homoeologous chromosomes, hexaploid wheat completes meiosis in a shorter period than diploid species with a much smaller genome. Therefore, during wheat meiosis, mechanisms additional to the classical model based on DNA sequence homology, must facilitate more efficient homologous recognition. One such mechanism could involve exploitation of differences in chromosome structure between homologues and homoeologues at the onset of meiosis. In turn, these chromatin changes, can be expected to be linked to transcriptional gene activity. In this study, we present an extensive analysis of a large RNA-Seq data derived from six different genotypes: wheat, wheat-rye hybrids and newly synthesized octoploid triticale, both in the presence and absence of the Ph1 locus. Plant material was collected at early prophase, at the transition leptotene-zygotene, when the telomere bouquet is forming and synapsis between homologues is beginning. The six genotypes exhibit different levels of synapsis and chromatin structure at this stage; therefore, recombination and consequently segregation, are also different. Unexpectedly, our study reveals that neither synapsis, whole genome duplication nor the absence of the Ph1 locus are associated with major changes in gene expression levels during early meiotic prophase. Overall wheat transcription at this meiotic stage is therefore highly resilient to such alterations, even in the presence of major chromatin structural changes. This suggests that post-transcriptional and post-translational processes are likely to be more important. Thus, further studies will be required to reveal whether these observations are specific to wheat meiosis, and whether there are significant changes in post-transcriptional and post-translational modifications in wheat and other polyploid species associated with their polyploidisation.

Exploiting the ZIP4 homologue within the wheat Ph1 locus has identified two lines exhibiting homoeologous crossover in wheat-wild relative hybrids

Despite possessing related ancestral genomes, hexaploid wheat behaves as a diploid during meiosis. The wheat Ph1 locus promotes accurate synapsis and crossover of homologous chromosomes. Interspecific hybrids between wheat and wild relatives are exploited by breeders to introgress important traits from wild relatives into wheat, although in hybrids between hexaploid wheat and wild relatives, which possess only homoeologues, crossovers do not take place during meiosis at metaphase I. However, in hybrids between Ph1 deletion mutants and wild relatives, crossovers do take place. A single Ph1 deletion (ph1b) mutant has been exploited for the last 40 years for this activity. We show here that selection of chemical induced mutant lines possessing mutations in TaZIP4-B2 exhibit high levels of homoeologous crossovers when crossed with a wild relative. Exploitation of Tazip4-B2 mutants rather than mutants with whole Ph1 locus deletions may improve introgression of wild relative chromosome segments into wheat. Such mutant lines may be more stable over multiple generations, as multivalents causing accumulation of chromosome translocations are less frequent.Key messageExploiting the ZIP4 homologue within the wheat Ph1 locus has identified two wheat mutants through a non-GM route, which can be exploited as an alternative to the Chinese Spring ph1b mutant in wheat introgression strategies.