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The Arabidopsis HOP2 gene has a role in preventing illegitimate connections between nonhomologous chromosome regions

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Abstract

Meiotic homologous chromosomes synapse and undergo crossing over (CO). In many eukaryotes, both synapsis and crossing over require the induction of double stranded breaks (DSBs) and subsequent repair via homologous recombination. In these organisms, two key proteins are recombinases RAD51 and DMC1. Recombinase-modulators HOP2 and MND1 assist RAD51 and DMC1 and also are required for synapsis and CO. We have investigated the hop2-1 phenotype in Arabidopsis during the segregation stages of both meiosis and mitosis. Despite a general lack of synapsis during prophase I, we observed extensive, stable interconnections between nonhomologous chromosomes in diploid hop2-1 nuclei in first and second meiotic divisions. Using γH2Ax as a marker of unrepaired DSBs, we detected γH2AX foci from leptotene through early pachytene but saw no foci from mid-pachytene onward. We conclude that the bridges seen from metaphase I onward are due to mis-repaired DSBs, not unrepaired ones. Examining haploids, we found that wild type haploids produce only univalents, but hop2-1 haploids like hop2-1 diploids have illegitimate connections stable enough to produce bridged chromosomes during segregation. Our results suggest that HOP2 has a significant active role in preventing repairs that use nonhomologous chromosomes during meiosis. We also found evidence that HOP2 plays a role in preventing illegitimate repair of radiation-induced DSBs in rapidly dividing petal cells. We conclude that HOP2 in Arabidopsis plays both a positive role in promoting synapsis and a separable role in preventing DSB repair using nonhomologous chromosomes.

Significance statement

The fidelity of homologous recombination (HR) during meiosis is essential to the production of viable gametes and for maintaining genome integrity in vegetative cells. HOP2 is an important protein for accurate meiotic HR in plants. We have found evidence of high levels of illegitimate repairs between nonhomologous chromosomes during meiosis and in irradiated petal cells in hop2-1 mutants, suggesting a role for HOP2 beyond its established role in synapsis and crossing over.

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Data availability

All data presented in this paper are found in either the Results section or Supplemental materials.

Code availability

NA

Abbreviations

CO:

Crossovers

Col:

Columbia

DSB :

Double strand break

DHJ:

Double Holliday Junctions

EdU:

5-Ethynyl-2′-deoxyuridine

FISH:

Flourescence in situ hybridization

HR:

Homologous eecombination

Ler :

Landsberg erecta

MI-AI:

Metaphase I-anaphase I interval

NHEJ:

Nonhomologous End joining

PSU:

Pistil stamen units

SC:

Synaptonemal complex

SDSA:

Synthesis-dependent strand annealing

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Acknowledgements

The authors appreciate the advice, sharing of equipment and/or technical assistance of Dr. Sohee Kang and Bruno Chue and colleagues in the Plant Science group in the Department of Biological Sciences at the University of Toronto Scarborough. We also acknowledge undergraduates Nick Garrido, Diane Hamdan and Ellie Kubisz who played small but important roles in this research. Special thanks to Dr. Patti Stronghill who did most of the EdU and H2AX data collection. We also wish to thank the reviewers of this manuscript who made several very helpful revision suggestions.

Funding

The research was funded by a grant from the Natural Sciences and Engineering Research Council of Canada (NSERC) to CAH and CDR, grant RGPGP‐2015‐00071, and a NSERC summer research award to YF-T.

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Authors and Affiliations

  1. University of Toronto Scarborough, Toronto, ON, Canada

    Yisell Farahani-Tafreshi, Chun Wei, Peilu Gan, Jenya Daradur, C. Daniel Riggs & Clare A. Hasenkampf

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  1. Yisell Farahani-Tafreshi
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  2. Chun Wei
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  3. Peilu Gan
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  4. Jenya Daradur
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  6. Clare A. Hasenkampf
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Contributions

CAH designed the broad strokes of the experiments and wrote the paper. CDR developed and supervised all molecular biology experiments and provided major editorial assistance. PG did the preliminary work on generating and analyzing haploid plants and assisted in processing inflorescences for the EdU labeling. YF-T refined the cytological protocols and did genotyping, karyotyping, and cytological analysis for the haploid experiments. CW developed protocols and collected all of the data for the petal irradiation experiments. JD performed genotyping and trained undergraduate personnel.

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Correspondence to Clare A. Hasenkampf.

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Farahani-Tafreshi, Y., Wei, C., Gan, P. et al. The Arabidopsis HOP2 gene has a role in preventing illegitimate connections between nonhomologous chromosome regions. Chromosome Res 30, 59–75 (2022). https://doi.org/10.1007/s10577-021-09681-2

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