The meiotic topoisomerase VI B subunit (MTOPVIB) is essential for meiotic DNA double-strand break formation in barley (Hordeum vulgare L.)

Steckenborn, Stefan; Cuacos, María; Ayoub, Mohammad A.; Chao, Feng; Schubert, Veit; Hoffie, Iris; Hensel, Goetz; Kumlehn, Jochen; Heckmann, Stefan

doi:10.1007/s00497-022-00444-5

Cited by 8 publications

(4 citation statements)

References 80 publications

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“…In most previous analyses of plant species where immunocytochemistry has been used to indirectly estimate meiotic DSBs, mutation of either SPO11‐1 , SPO11‐2 , or MTOPVIB resulted in a dramatic reduction in foci numbers implying a corresponding reduction in DSBs (Benyahya et al ., 2020 ; de Muyt et al ., 2007 ; Fayos et al ., 2020 ; Ku et al ., 2020 ; Li et al ., 2022 ; Sanchez‐Moran et al ., 2007 ; Steckenborn et al ., 2022 ). For example, in maize and rice, mutation of SPO11‐1 was reported to result in the reduction of DSBs by ~97% and ~83%, respectively (Fayos et al ., 2020 ; Ku et al ., 2020 ).…”

Section: Discussionmentioning

confidence: 99%

“…The catalytically active tyrosine residues of both SPO11‐1 and SPO11‐2 are required for break formation (Hartung et al ., 2007 ). Given the high degree of functional and sequence conservation of plant SPO11 proteins, it seems likely that wheat SPO11‐1 and SPO11‐2 similarly form a catalytic heterodimer within a heterotetrameric DSB complex, especially as the MTOPVIB protein is also functionally conserved in cereals (Jing et al ., 2020 ; Steckenborn et al ., 2022 ; Xue et al ., 2016 ).…”

Section: Discussionmentioning

confidence: 99%

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Identification, characterization, and rescue of CRISPR/Cas9 generated wheat SPO11‐1 mutants

Hyde

Osman

Winfield

et al. 2022

Plant Biotechnology Journal

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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. The 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, 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. Characterization 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 the 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.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Identification, characterization, and rescue of CRISPR/Cas9 generated wheat SPO11‐1 mutants

Hyde

Osman

Winfield

et al. 2022

Plant Biotechnology Journal

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“…Somatic cmf7 mutations, induced in virus-infected plants, can be transmitted to subsequent generations resulting in albino or variegated plants defective for CMF7 . In addition, somatic mutations were induced in three meiosis-related genes in barley: ASY1 , encoding for a meiotic chromosome axis-localized HORMA domain protein ( Armstrong et al., 2002 ; Steckenborn et al., 2023 ), MUS81 , encoding for a DNA structure-selective endonuclease involved in the formation of meiotic class II crossover (CO) ( Hartung et al., 2006 ; Berchowitz et al., 2007 ; Higgins et al., 2008 ) as well as ZYP1 , encoding for a transverse filament protein of the synaptonemal complex (SC) ( Barakate et al., 2014 ; Steckenborn et al., 2023 ). Hence, BSMVIGE enables rapid somatic and heritable targeted gene editing in barley.…”

Section: Introductionmentioning

confidence: 99%

Barley stripe mosaic virus-mediated somatic and heritable gene editing in barley (Hordeum vulgare L.)

Tamilselvan-Nattar-Amutha,

Hiekel,

Hartmann

et al. 2023

Front. Plant Sci.

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Genome editing strategies in barley (Hordeum vulgare L.) typically rely on Agrobacterium-mediated genetic transformation for the delivery of required genetic reagents involving tissue culture techniques. These approaches are genotype-dependent, time-consuming, and labor-intensive, which hampers rapid genome editing in barley. More recently, plant RNA viruses have been engineered to transiently express short guide RNAs facilitating CRISPR/Cas9-based targeted genome editing in plants that constitutively express Cas9. Here, we explored virus-induced genome editing (VIGE) based on barley stripe mosaic virus (BSMV) in Cas9-transgenic barley. Somatic and heritable editing in the ALBOSTRIANS gene (CMF7) resulting in albino/variegated chloroplast-defective barley mutants is shown. In addition, somatic editing in meiosis-related candidate genes in barley encoding ASY1 (an axis-localized HORMA domain protein), MUS81 (a DNA structure-selective endonuclease), and ZYP1 (a transverse filament protein of the synaptonemal complex) was achieved. Hence, the presented VIGE approach using BSMV enables rapid somatic and also heritable targeted gene editing in barley.

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“…Using this technology, target sequence‐specified guide RNAs (gRNAs) are capable of precisely directing Cas endonucleases to predetermined positions in any candidate gene of choice, where the endonucleases induce DNA double‐strand breaks whose error‐prone repair can cause random mutations at the mended site. In barley, too, Cas endonuclease technology has been applied in some studies to validate gene functions or to introduce advantageous traits without any further modification of the genetic background (Gerasimova et al ., 2020 ; Hisano et al ., 2022 ; Hoffie et al ., 2021 ; Li et al ., 2019 ; Steckenborn et al ., 2022 ). Since all potential rym11 donors represent rather exotic germplasm for winter barley breeding, the site‐directed generation of rym11 alleles is considered as being a particularly useful approach towards the development of virus‐resistant varieties.…”

Section: Introductionmentioning

confidence: 99%

Novel resistance to the BymovirusBaMMVestablished by targeted mutagenesis of thePDIL5‐1susceptibility gene in barley

Hoffie

Perović

Habekuß

et al. 2022

Plant Biotechnology Journal

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The Potyviridae are the largest family of plant-pathogenic viruses. Members of this family are the soil-borne bymoviruses barley yellow mosaic virus (BaYMV) and barley mild mosaic virus (BaMMV), which, upon infection of young winter barley seedlings in autumn, can cause yield losses as high as 50%. Resistance breeding plays a major role in coping with these pathogens. However, some viral strains have overcome the most widely used resistance. Thus, there is a need for novel sources of resistance. In ancient landraces and wild relatives of cultivated barley, alleles of the susceptibility factor PROTEIN DISULFIDE ISOMERASE LIKE 5-1 (PDIL5-1) were identified to confer resistance to all known strains of BaYMV and BaMMV. Although the gene is highly conserved throughout all eukaryotes, barley is thus far the only species for which PDIL5-1-based virus resistance has been reported. Whereas introgression by crossing to the European winter barley breeding pool is tedious, time-consuming and additionally associated with unwanted linkage drag, the present study exemplifies an approach to targeted mutagenesis of two barley cultivars employing CRISPR-associated endonuclease technology to induce site-directed mutations similar to those described for PDIL5-1 alleles that render certain landraces resistant. Homozygous primary mutants were produced in winter barley, and transgene-free homozygous M 2 mutants were produced in spring barley. A variety of mutants carrying novel PDIL5-1 alleles were mechanically inoculated with BaMMV, by which all frameshift mutations and certain in-frame mutations were demonstrated to confer resistance to this virus. Under greenhouse conditions, virus-resistant mutants showed no adverse effects in terms of growth and yield.

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The meiotic topoisomerase VI B subunit (MTOPVIB) is essential for meiotic DNA double-strand break formation in barley (Hordeum vulgare L.)

Cited by 8 publications

References 80 publications

Identification, characterization, and rescue of CRISPR/Cas9 generated wheat SPO11‐1 mutants

Identification, characterization, and rescue of CRISPR/Cas9 generated wheat SPO11‐1 mutants

Barley stripe mosaic virus-mediated somatic and heritable gene editing in barley (Hordeum vulgare L.)

Novel resistance to the BymovirusBaMMVestablished by targeted mutagenesis of thePDIL5‐1susceptibility gene in barley

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