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Wheat is a major cereal crop that possesses a large allopolyploid genome formed through hybridisation of tetraploid and diploid progenitors. During meiosis, crossovers (COs) are constrained in number to 1–3 per chromosome pair that are predominantly located towards the chromosome ends. This reduces the probability of advantageous traits recombining onto the same chromosome, thus limiting breeding. Therefore, understanding the underlying factors controlling meiotic recombination may provide strategies to unlock the genetic potential in wheat. In this mini-review, we will discuss the factors associated with restricted CO formation in wheat, such as timing of meiotic events, chromatin organisation, pre-meiotic DNA replication and dosage of CO genes, as a means to modulate recombination.

Our presentation will discuss how our team of Indigenous and non-Indigenous developed a unique Indigenous research methodology to increase understanding of the digital inclusion challenges faced by low income Indigenous families living on Mornington Island, a remote Aboriginal community in remote Queensland, Australia. We discuss our methodology and findings of how low income Mornington Island Aboriginal families use the internet to access government and social services, education, e-health and e-commerce. This project employs Indigenous methodologies including yarning, participant photography, and co-design workshops to inform policies, practices and programs. Our results are interpreted within the growing body of decolonizing methodologies and Indigenous research, methods and methodologies. Informed by Martin’s (2009) Relatedness Theory, our data is considered through Aboriginal ways of being, doing and knowing, with Aboriginal axiology, epistemology and ontology forming a window through which we observe, analyse and interpret data. In this presentation, we will discuss how we as a blended research team: • facilitated Indigenous involvement in the research (starting with the development of the research concept and design) • engaged strongly with with Indigenous research and literature, both academic and community-based to interpret data and findings • engaged with and developed Indigenous methodologies to ensure the effectiveness and appropriateness of this research and • represented Indigenous authors and voices in analysis and communication of research results

This article outlines and demonstrates how multi-platform issue mapping can be used to investigate the impact of scholarly contributions to public debates, using digital methods and data drawn from...

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Meiosis generates genetic variation through homologous recombination (HR) that is harnessed during breeding. HR occurs in the context of meiotic chromosome axes and the synaptonemal complex. To study the role of axis remodelling in crossover (CO) formation in a crop species, we characterized mutants of the axis-associated protein ASY1 and the axis-remodelling protein PCH2 in Brassica rapa. asy1 plants form meiotic chromosome axes that fail to synapse. CO formation is almost abolished, and residual chiasmata are proportionally enriched in terminal chromosome regions, particularly in the nucleolar organizing region (NOR)-carrying chromosome arm. pch2 plants show impaired ASY1 loading and remodelling, consequently achieving only partial synapsis, which leads to reduced CO formation and loss of the obligatory CO. PCH2-independent chiasmata are proportionally enriched towards distal chromosome regions. Similarly, in Arabidopsis pch2, COs are increased towards telomeric regions at the expense of (peri-) centromeric COs compared with the wild type. Taken together, in B. rapa, axis formation and remodelling are critical for meiotic fidelity including synapsis and CO formation, and in asy1 and pch2 CO distributions are altered. While asy1 plants are sterile, pch2 plants are semi-sterile and thus PCH2 could be an interesting target for breeding programmes.

Meiotic recombination generates genetic variation and provides physical links between homologous chromosomes (crossovers) essential for accurate segregation. In cereals the distribution of crossovers, cytologically evident as chiasmata, is biased toward the distal regions of chromosomes. This creates a bottleneck for plant breeders in the development of varieties with improved agronomic traits, as genes situated in the interstitial and centromere proximal regions of chromosomes rarely recombine. Recent advances in wheat genomics and genome engineering combined with well-developed wheat cytogenetics offer new opportunities to manipulate recombination and unlock genetic variation. As a basis for these investigations we have carried out a detailed analysis of meiotic progression in hexaploid wheat (Triticum aestivum) using immunolocalization of chromosome axis, synaptonemal complex and recombination proteins. 5-Bromo-2′-deoxyuridine (BrdU) labeling was used to determine the chronology of key events in relation to DNA replication. Axis morphogenesis, synapsis and recombination initiation were found to be spatio-temporally coordinated, beginning in the gene-dense distal chromosomal regions and later occurring in the interstitial/proximal regions. Moreover, meiotic progression in the distal regions was coordinated with the conserved chromatin cycles that are a feature of meiosis. This mirroring of the chiasma bias was also evident in the distribution of the gene-associated histone marks, H3K4me3 and H3K27me3; the repeat-associated mark, H3K27me1; and H3K9me3. We believe that this study provides a cytogenetic framework for functional studies and ongoing initiatives to manipulate recombination in the wheat genome.

During meiosis chromosomes undergo DNA double-strand breaks (DSBs) that can be repaired using a homolog to produce crossovers, which creates genetic diversity. Meiotic recombination occurs coincident with homolog pairing and polymerization of the meiotic axis and synaptonemal complex (SC). REC8-cohesin is required to connect chromosomes to the axis and to organize axis polymerization. However, control of REC8 loading along chromosomes, in relation to chromatin, transcription and recombination, is not yet fully understood. Therefore, we performed REC8 ChIP-seq in Arabidopsis, which revealed strong enrichment in centromeric heterochromatin. REC8 abundance correlates with suppression of meiotic DSBs and crossovers, despite axis loading of SPO11-1 in these regions. Loss of the heterochromatic marks H3K9me2 and non-CG DNA methylation in kyp/suvh4 suvh5 suvh6 mutants causes remodeling of REC8 and gain of meiotic recombination locally in repeated sequences, although centromere cohesion is maintained. In the chromosome arms, REC8 is enriched within gene bodies, exons and GC-rich sequences, and anti-correlates with transcription. Highest REC8 occupancy occurred in facultatively silent, H3K27me3-modified genes. Using immunocytology we show that axis polycomplexes form in rec8 mutants that recruit recombination foci with altered stoichiometry, leading to catastrophic non-homologous recombination. Therefore, REC8 plays a key role organizing meiotic chromosome architecture and promoting high-fidelity interhomolog recombination. Despite this pro-recombination role, local REC8 enrichment associates with DSB repression at the fine scale, which is consistent with the tethered-loop/axis model. Coincident with its organizational role during meiosis, REC8-cohesin occupancy along the chromosomes is shaped by multiple chromatin states and transcription.

International audience; During the leptotene stage of prophase I of meiosis, chromatids become organized into a linear looped array via a protein axis that forms along the loop bases. Establishment of the axis is essential for the subsequent synapsis of the homologous chromosome pairs and the progression of recombination to form genetic crossovers. Here, we describe ASYNAPTIC4 (ASY4), a meiotic axis protein in Arabidopsis (Arabidopsis thaliana). ASY4 is a small coiled-coil protein that exhibits limited sequence similarity with the carboxyl-terminal region of the axis protein ASY3. We used enhanced yellow fluorescent protein-tagged ASY4 to show that ASY4 localizes to the chromosome axis throughout prophase I. Bimolecular fluorescence complementation revealed that ASY4 interacts with ASY1 and ASY3, and yeast two-hybrid analysis confirmed a direct interaction between ASY4 and ASY3. Mutants lacking full-length ASY4 exhibited defective axis formation and were unable to complete synapsis. Although the initiation of recombination appeared to be unaffected in the asy4 mutant, the number of crossovers was reduced significantly, and crossovers tended to group in the distal parts of the chromosomes. We conclude that ASY4 is required for normal axis and crossover formation. Furthermore, our data suggest that ASY3/ASY4 are the functional homologs of the mammalian SYCP2/SYCP3 axial components.

The movement of chromosomes during meiosis involves location of their telomeres at the inner surface of the nuclear envelope. Sad1/UNC-84 (SUN) domain proteins are inner nuclear envelope proteins that are part of complexes linking cytoskeletal elements with the nucleoskeleton, connecting telomeres to the force-generating mechanism in the cytoplasm. These proteins play a conserved role in chromosome dynamics in eukaryotes. Homologues of SUN domain proteins have been identified in several plant species. In Arabidopsis thaliana, two proteins that interact with each other, named AtSUN1 and AtSUN2, have been identified. Immunolocalization using antibodies against AtSUN1 and AtSUN2 proteins revealed that they were associated with the nuclear envelope during meiotic prophase I. Analysis of the double mutant Atsun1-1 Atsun2-2 has revealed severe meiotic defects, namely a delay in the progression of meiosis, absence of full synapsis, the presence of unresolved interlock-like structures, and a reduction in the mean cell chiasma frequency. We propose that in Arabidopsis thaliana, overlapping functions of SUN1 and SUN2 ensure normal meiotic recombination and synapsis.

RecQ helicases are a conserved group of proteins with a role in the maintenance of genome integrity. In Saccharomyces cerevisiae (budding yeast), meiotic recombination is increased in the absence of the RecQ helicase Sgs1. Here we investigated the potential meiotic role of the Sgs1 homologue AtRECQ4A and the closely related AtRECQ4B. Both proteins have been shown to function during recombination in somatic cells, but so far their meiotic role has not been investigated. Both AtRECQ4A and AtRECQ4B were expressed in reproductive tissues. Although immunolocalization studies showed that AtRECQ4A associates with recombination intermediates, we found no evidence that its loss or that of AtRECQ4B had a significant effect on meiotic cross-overs, suggesting functional redundancy with other RECQ family members. Nevertheless, pollen viability decreased in Atrecq4A, resulting in a reduction in fertility, although this was not the case in Atrecq4B. Cytological analysis revealed chromatin bridges between the telomeres of non-homologous chromosomes in Atrecq4A at metaphase I, in some instances accompanied by chromosome fragmentation at anaphase I. The bridges required telomeric repeats and were dependent on meiotic recombination. Immunolocalization confirmed the association of AtRECQ4A with the telomeres during prophase I, which we propose enables dissolution of recombination-dependent telomeric associations. Thus, this study has identified a hitherto unknown role for a member of the RECQ helicase family during meiosis that contributes to the maintenance of chromosome integrity. As telomere structure is generally conserved, it seems likely that these associations may arise during meiosis in other species, where they must also be removed.

The ends of linear eukaryotic chromosomes are protected by the telomere, a specialised nucleoprotein complex. The primary role of the telomere is to protect the chromosome ends from being degraded or recognised and processed as double strand breaks. Additionally, the telomeres are involved in interphase chromosome organisation and also in chromosome pairing, synapsis and movement during meiotic prophase. The main emphasis of this review is concerned with the distribution and dynamics of the telomeres in the somatic cell and meiocytes of plants, focusing on the model plant Arabidopsis thaliana. In Arabidopsis the telomeres are organised around the nucleolus in both the somatic and meiotic interphase. One of the outstanding questions in meiosis is how homologous chromosomes pair (align) and synapse during meiotic prophase. Recent attention has been paid to the bouquet formation, a nearly universal event, during which the telomeres cluster on the nuclear membrane in early prophase. It has been suggested that because the telomeres are in close proximity this would enhance their pairing and subsequent synapsis of the homologues. In Arabidopsis we observe that the telomeres are paired homologously in early meiosis whilst still arranged around the nucleolus. They are moved to the nuclear membrane preceding synapsis and reveal only a loose clustering, which may represent a transient bouquet. On completion of synapsis the paired telomeres are dispersed and remain attached to the nuclear membrane until diplotene when they dissociate from the nuclear membrane. We also discuss the prospects for live imaging of the telomeres in Arabidopsis.

Meiosis is a fundamental and evolutionarily conserved process that is central to the life cycles of all sexually reproducing eukaryotes. An understanding of this process is critical to furthering research on reproduction, fertility, genetics and breeding. Plants have been used extensively in cytogenetic studies of meiosis during the last century. Until recently, our knowledge of the molecular and functional aspects of meiosis has emerged from the study of non-plant model organisms, especially budding yeast. However, the emergence of Arabidopsis thaliana as the model organism for plant molecular biology and genetics has enabled significant progress in the characterisation of key genes and proteins controlling plant meiosis. The development of molecular and cytological techniques in Arabidopsis, besides allowing investigation of the more conserved aspects of meiosis, are also providing insights into features of this complex process which may vary between organisms.This review highlights an example of this recent progress by focussing on ASY1, a meiosis-specific Arabidopsis protein which shares some similarity with the N-terminus region of the yeast axial core-associated protein, HOP1, a component of a multiprotein complex which acts as a meiosis-specific barrier to sister-chromatid repair in budding yeast. In the absence of ASY1, synapsis is interrupted and chiasma formation is dramatically reduced. ASY1 protein is initially detected during early meiotic G2 as numerous foci distributed over the chromatin. As G2 progresses the signal appears to be increasingly continuous and is closely associated with the axial elements. State-of-the-art cytogenetic techniques have revealed that initiation of recombination is synchronised with the formation of the chromosome axis. Furthermore, in the context of the developing chromosome axes, ASY1 plays a crucial role in co-ordinating the activity of a key member of the homologous recombination machinery, AtDMC1.

In higher plants, sexual reproduction involves interactions between pollen and pistil. A key mechanism to prevent inbreeding is self-incompatibility through rejection of incompatible ('self') pollen1. In Papaver rhoeas, S proteins encoded by the stigma interact with incompatible pollen, triggering a Ca2+-dependent signalling network2, 3, 4, 5 resulting in pollen tube inhibition and programmed cell death6. The cytosolic phosphoprotein p26.1, which has been identified in incompatible pollen, shows rapid, self-incompatibility-induced Ca2+-dependent hyperphosphorylation in vivo3. Here we show that p26.1 comprises two proteins, Pr-p26.1a and Pr-p26.1b, which are soluble inorganic pyrophosphatases (sPPases). These proteins have classic Mg2+-dependent sPPase activity, which is inhibited by Ca2+, and unexpectedly can be phosphorylated in vitro. We show that phosphorylation inhibits sPPase activity, establishing a previously unknown mechanism for regulating eukaryotic sPPases. Reduced sPPase activity is predicted to result in the inhibition of many biosynthetic pathways, suggesting that there may be additional mechanisms of self-incompatibility-mediated pollen tube inhibition. We provide evidence that sPPases are required for growth and that self-incompatibility results in an increase in inorganic pyrophosphate, implying a functional role for Pr-p26.1.

The application of proteomics techniques to the study of plant meiosis has the potential to make a valuable contribution to our understanding of the molecular events underpinning meiotic processes. Here we describe the preparation of meiotic protein complexes from Arabidopsis thaliana and its close crop relative, Brassica oleracea, by co-immunoprecipitation for in-solution analysis by tandem mass spectrometry (MS/MS). Early results using these techniques have proved encouraging, enabling the identification of candidate AtASY1-interacting proteins in A. thaliana and providing evidence of an in planta interaction between BoASY1 and BoASY3 in B. oleracea. The detection of phospho-modified peptides of BoASY1 and BoASY3 suggests that this approach may be useful for studying meiotic protein modification events.

Advances in the molecular biology and genetics of Arabidopsis thaliana have led to it becoming an important model for the analysis of meiosis in plants. Cytogenetic investigations are pivotal to meiotic studies and a number of technological improvements for Arabidopsis cytology have provided a range of tools to investigate chromosome behavior during meiosis (Jones et al. Chromosome Res 11:205-215, 2003). This chapter contains a detailed description of an immunological technique currently used in our lab for the preparation of meiotic chromosomes for immunolocalization.

Following successful completion of the Brassica rapa sequencing project, the next step is to investigate functions of individual genes/proteins. For Arabidopsis thaliana, large amounts of protein-protein interaction (PPI) data are available from the major PPI databases (DBs). It is known that Brassica crop species are closely related to A. thaliana. This provides an opportunity to infer the B. rapa interactome using PPI data available from A. thaliana. In this paper, we present an inferred B. rapa interactome that is based on the A. thaliana PPI data from two resources: (i) A. thaliana PPI data from three major DBs, BioGRID, IntAct, and TAIR. (ii) ortholog-based A. thaliana PPI predictions. Linking between B. rapa and A. thaliana was accomplished in three complementary ways: (i) ortholog predictions, (ii) identification of gene duplication based on synteny and collinearity, and (iii) BLAST sequence similarity search. A complementary approach was also applied, which used known/predicted domain-domain interaction data. Specifically, since the two species are closely related, we used PPI data from A. thaliana to predict interacting domains that might be conserved between the two species. The predicted interactome was investigated for the component that contains known A. thaliana meiotic proteins to demonstrate its usability.

Advances in the molecular biology and genetics of Arabidopsis thaliana have led to it becoming an important model for the analysis of meiosis in plants. Cytogenetic investigations are pivotal to meiotic studies and a number of technological improvements for Arabidopsis cytology have provided a range of tools to investigate chromosome behavior during meiosis (Jones et al. Chromosome Res 11:205-215, 2003). This chapter contains a detailed description of an immunological technique currently used in our lab for the preparation of meiotic chromosomes for immunolocalization.

The application of proteomics techniques to the study of plant meiosis has the potential to make a valuable contribution to our understanding of the molecular events underpinning meiotic processes. Here we describe the preparation of meiotic protein complexes from Arabidopsis thaliana and its close crop relative, Brassica oleracea, by co-immunoprecipitation for in-solution analysis by tandem mass spectrometry (MS/MS). Early results using these techniques have proved encouraging, enabling the identification of candidate AtASY1-interacting proteins in A. thaliana and providing evidence of an in planta interaction between BoASY1 and BoASY3 in B. oleracea. The detection of phospho-modified peptides of BoASY1 and BoASY3 suggests that this approach may be useful for studying meiotic protein modification events.

Contents Summary 523 I. Introduction 524 II. The meiotic pathway: a brief overview 524 III. Homologous chromosome pairing and movement during prophase I 525 IV. Meiotic DNA double-strand break formation 526 V. Processing of DNA double-strand breaks 528 VI. Strand exchange: the role of the RecA homologues and their accessory proteins 529 VII. Promotion of stable strand exchange 532 VIII. Pathways to crossover formation 532 IX. The class I pathway of meiotic recombination 533 X. The class II pathway of meiotic recombination 536 XI. Holliday junction (Hj) resolution 537 XII. Noncrossover pathways and the crossover/ noncrossover decision 537 XIII. Conclusions 538 Acknowledgements 538 References 538 Summary Meiosis is a central feature of sexual reproduction. Studies in plants have made and continue to make an important contribution to fundamental research aimed at the understanding of this complex process. Moreover, homologous recombination during meiosis provides the basis for plant breeders to create new varieties of crops. The increasing global demand for food, combined with the challenges from climate change, will require sustained efforts in crop improvement. An understanding of the factors that control meiotic recombination has the potential to make an important contribution to this challenge by providing the breeder with the means to make fuller use of the genetic variability that is available within crop species. Cytogenetic studies in plants have provided considerable insights into chromosome organization and behaviour during meiosis. More recently, studies, predominantly in Arabidopsis thaliana, are providing important insights into the genes and proteins that are required for crossover formation during plant meiosis. As a result, substantial progress in the understanding of the molecular mechanisms that underpin meiosis in plants has begun to emerge. This article summarizes current progress in the understanding of meiotic recombination and its control in Arabidopsis. We also assess the relationship between meiotic recombination in Arabidopsis and other eukaryotes, highlighting areas of close similarity and apparent differences.

RecQ helicases are a conserved group of proteins with a role in the maintenance of genome integrity. In Saccharomyces cerevisiae (budding yeast), meiotic recombination is increased in the absence of the RecQ helicase Sgs1. Here we investigated the potential meiotic role of the Sgs1 homologue AtRECQ4A and the closely related AtRECQ4B. Both proteins have been shown to function during recombination in somatic cells, but so far their meiotic role has not been investigated. Both AtRECQ4A and AtRECQ4B were expressed in reproductive tissues. Although immunolocalization studies showed that AtRECQ4A associates with recombination intermediates, we found no evidence that its loss or that of AtRECQ4B had a significant effect on meiotic cross-overs, suggesting functional redundancy with other RECQ family members. Nevertheless, pollen viability decreased in Atrecq4A, resulting in a reduction in fertility, although this was not the case in Atrecq4B. Cytological analysis revealed chromatin bridges between the telomeres of non-homologous chromosomes in Atrecq4A at metaphase I, in some instances accompanied by chromosome fragmentation at anaphase I. The bridges required telomeric repeats and were dependent on meiotic recombination. Immunolocalization confirmed the association of AtRECQ4A with the telomeres during prophase I, which we propose enables dissolution of recombination-dependent telomeric associations. Thus, this study has identified a hitherto unknown role for a member of the RECQ helicase family during meiosis that contributes to the maintenance of chromosome integrity. As telomere structure is generally conserved, it seems likely that these associations may arise during meiosis in other species, where they must also be removed.

Higher plants produce seed through pollination, using specific interactions between pollen and pistil. Self-incompatibility is an important mechanism used in many species to prevent inbreeding; it is controlled by a multi-allelic S locus1, 2. ‘Self’ (incompatible) pollen is discriminated from ‘non-self’ (compatible) pollen by interaction of pollen and pistil S locus components, and is subsequently inhibited. In Papaver rhoeas, the pistil S locus product is a small protein that interacts with incompatible pollen, triggering a Ca2+-dependent signalling network, resulting in pollen inhibition and programmed cell death3, 4, 5, 6, 7. Here we have cloned three alleles of a highly polymorphic pollen-expressed gene, PrpS (Papaver rhoeas pollen S), from Papaver and provide evidence that this encodes the pollen S locus determinant. PrpS is a single-copy gene linked to the pistil S gene (currently called S, but referred to hereafter as PrsS for Papaver rhoeas stigma S determinant). Sequence analysis indicates that PrsS and PrpS are equally ancient and probably co-evolved. PrpS encodes a novel ~20-kDa protein. Consistent with predictions that it is a transmembrane protein, PrpS is associated with the plasma membrane. We show that a predicted extracellular loop segment of PrpS interacts with PrsS and, using PrpS antisense oligonucleotides, we demonstrate that PrpS is involved in S-specific inhibition of incompatible pollen. Identification of PrpS represents a major advance in our understanding of the Papaver self-incompatibility system. As a novel cell–cell recognition determinant it contributes to the available information concerning the origins and evolution of cell–cell recognition systems involved in discrimination between self and non-self, which also include histocompatibility systems in primitive chordates and vertebrates.

Replication protein A (RPA) is involved in many aspects of DNA metabolism including meiotic recombination. Many species possess a single RPA1 gene but Arabidopsis possesses five RPA1 paralogues. This feature has enabled us to gain further insight into the meiotic role of RPA1. Proteomic analysis implicated one of the AtRPA1 family (AtRPA1a) in meiosis. Immunofluorescence studies confirmed that AtRPA1a is associated with meiotic chromosomes from leptotene through to early pachytene. Analysis of an Atrpa1a mutant revealed that AtRPA1a is not essential at early stages in the recombination pathway. DNA double-strand breaks are repaired in Atrpa1a, but the mutant is defective in the formation of crossovers, exhibiting a 60% reduction in chiasma frequency. Consistent with this, localization of recombination proteins AtRAD51 and AtMSH4 appears normal, whereas the numbers of AtMLH1 and AtMLH3 foci at pachytene are significantly reduced. This suggests that the defect in Atrpa1a is manifested at the stage of second-end capture. Analysis of Atrpa1a/Atmsh4 and Atrpa1a/Atmlh3 double mutants indicates that loss of AtRPA1a predominantly affects the formation of class I, interference-dependent crossovers.

Nature 459, 992–995 (2009); doi: 10.1038/nature08027 Recently, it has come to our attention that in the left panel of Fig. 2b of this Letter, the lanes labelled S2S4 and S6S17 were duplicated. We have reviewed the original data. It seems likely that a duplicated part of the blot was placed over laneS6S17 to aid alignment of molecular mass markers and inadvertently left there.

Immunocytochemistry reveals that the Arabidopsis mismatch repair proteins AtMSH4, AtMLH3 and AtMLH1 are expressed during prophase I of meiosis. Expression of AtMSH4 precedes AtMLH3 and AtMLH1 which co-localize as foci during pachytene. Co-localization between AtMSH4 and AtMLH3 occurs, but appears transient. AtMLH3 foci are not detected in an Atmsh4 mutant. However, localization of AtMSH4 is unaffected in Atmlh3, suggesting that recombination may proceed to dHj (double Holliday junction) formation. Mean chiasma frequency in Atmsh4 is reduced to 1.55 compared with 9.86 in wild-type. In contrast with wild-type, the distribution of residual crossovers in Atmsh4 closely fits a Poisson distribution. This is consistent with a two-pathway model for meiotic crossing-over whereby most crossovers occur via an AtMSH4-dependent pathway that is subject to interference, with the remaining crossovers arising via an interference-independent pathway. Loss of AtMLH3 results in an approx. 60% reduction in crossovers. Results suggest that dHj resolution can occur, but in contrast with wild-type where most or all dHjs are directed to form crossovers, the outcome is biased in favour of a non-crossover outcome. The results are compatible with a model whereby the MutL complex maintains or imposes a dHj conformation that ensures crossover formation.

With respect to history, plants have provided an ideal system for cytogenetical analysis of the synaptonemal complex (SC). However, until recently, the identification of the genes that encode the SC in plants has proved elusive. In recent years, Arabidopsis thaliana was developed as a model system for plant meiosis research. As a result, there was substantial progress in the isolation of meiotic genes and this has recently led to the isolation of the first plant SC gene, ZYP1. The ZYP1 gene encodes a transverse filament (TF) protein that is predicted to have structural similarity to TF proteins found in other organisms. Analysis of plants deficient in ZYP1 expression has provided important insights into the function of the SC in plants. Loss of ZYP1 has only a limited effect on the overall level of recombination. However, it is associated with extensive nonhomologous recombination leading to multivalent formation at metaphase I. This phenomenon was not previously reported in other organisms. It is important to note that cytological analysis of the ZYP1 deficient lines indicates that SC formation is not required for the imposition of crossover interference.

Mitogen-activated protein kinases (MAPKs) operate downstream of receptor–ligand interactions, playing a pivotal role in responses to extracellular signals. The self-incompatibility (SI) response in Papaver rhoeas L. triggers a Ca2+-dependent signalling cascade resulting in inhibition of incompatible pollen. We have investigated the possible involvement of MAPKs in SI. We report the enhanced activation of a 56 kDa protein kinase (p56) in SI-induced pollen and provide evidence that p56 has MAPK activity. This provides an important advance in our understanding of the SI response. We believe this is the first direct biochemical demonstration of activation of a MAPK during SI.

Sexual reproduction in flowering plants is controlled by recognition mechanisms involving the male gametophyte (the pollen) and the female sporophyte (the pistil). Self–incompatibility (SI) involves the recognition and rejection of self– or incompatible pollen by the pistil. In Papaver rhoeas , SI uses a Ca 2+ –based signalling cascade triggered by the S –protein, which is encoded by the stigmatic component of the S –locus. This results in the rapid inhibition of incompatible pollen tube growth. We have identified several targets of the SI signalling cascade, including protein kinases, the actin cytoskeleton and nuclear DNA. Here, we summarize progress made on currently funded projects in our laboratory investigating some of the components targeted by SI, comprising (i) the characterization of a pollen phosphoprotein (p26) that is rapidly phosphorylated upon an incompatible SI response; (ii) the identification and characterization of a pollen mitogen–activated protein kinase (p56), which exhibits enhanced activation during SI; (iii) characterizing components involved in the reorganization and depolymerization of the actin cytoskeleton during the SI response; and (iv) investigating whether the SI response involves a programmed cell death signalling cascade.

The hexaploid bread wheat genome comprises over 16 gigabases of sequence across 21 chromosomes. Meiotic crossovers are highly polarized along the chromosomes, with elevation in the gene-dense distal regions and suppression in the Gypsy retrotransposon-dense centromere-proximal regions. We profiled the genomic landscapes of the meiotic recombinase DMC1 and the chromosome axis protein ASY1 in wheat and investigated their relationships with crossovers, chromatin state, and genetic diversity. DMC1 and ASY1 chromatin immunoprecipitation followed by sequencing (ChIP-seq) revealed strong co-enrichment in the distal, crossover-active regions of the wheat chromosomes. Distal ChIP-seq enrichment is consistent with spatiotemporally biased cytological immunolocalization of DMC1 and ASY1 close to the telomeres during meiotic prophase I. DMC1 and ASY1 ChIP-seq peaks show significant overlap with genes and transposable elements in the Mariner and Mutator superfamilies. However, DMC1 and ASY1 ChIP-seq peaks were detected along the length of each chromosome, including in low-crossover regions. At the fine scale, crossover elevation at DMC1 and ASY1 peaks and genes correlates with enrichment of the Polycomb histone modification H3K27me3. This indicates a role for facultative heterochromatin, coincident with high DMC1 and ASY1, in promoting crossovers in wheat and is reflected in distalized H3K27me3 enrichment observed via ChIP-seq and immunocytology. Genes with elevated crossover rates and high DMC1 and ASY1 ChIP-seq signals are overrepresented for defense-response and immunity annotations, have higher sequence polymorphism, and exhibit signatures of selection. Our findings are consistent with meiotic recombination promoting genetic diversity, shaping host–pathogen co-evolution, and accelerating adaptation by increasing the efficiency of selection.

Increasing crop yields through plant breeding is time consuming and laborious, with the generation of novel combinations of alleles being limited by chromosomal linkage blocks and linkage-drag. Meiotic recombination is essential to create novel genetic variation via the reshuffling of parental alleles. Exchange of genetic information between homologous chromosomes occurs at crossover (CO) sites but CO frequency is often low and unevenly distributed. This bias creates the problem of linkage-drag in recombination 'cold' regions, where undesirable variation remains linked to useful traits. In plants, programmed meiosis-specific DNA double-strand breaks (DSBs), catalysed by the SPO11 complex, initiate the recombination pathway, although only ~5% result in the formation of COs. To study the role of SPO11-1 in wheat meiosis, and as a prelude to manipulation, we used CRISPR/Cas9 to generate edits in all three SPO11-1 homoeologues of hexaploid wheat. Characterisation of progeny lines shows plants deficient in all six SPO11-1 copies fail to undergo chromosome synapsis, lack COs and are sterile. In contrast, lines carrying a single copy of any one of the three wild-type homoeologues are phenotypically indistinguishable from unedited plants both in terms of vegetative growth and fertility. However, cytogenetic analysis of the edited plants suggests that homoeologues differ in their ability to generate COs and in the dynamics of synapsis. In addition, we show that transformation of wheat mutants carrying six edited copies of SPO11-1 with the TaSPO11-1B gene, restores synapsis, CO formation and fertility and hence opens a route to modifying recombination in this agronomically important crop.