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Heterologous synapsis in C. elegans is regulated by meiotic double-strand breaks and crossovers

Abstract

Alignment of the parental chromosomes during meiotic prophase is key to the formation of genetic exchanges, or crossovers, and consequently to the successful production of gametes. In almost all studied organisms, alignment involves synapsis: the assembly of a conserved inter-chromosomal interface called the synaptonemal complex (SC). While the SC usually synapses homologous sequences, it can assemble between heterologous sequences. However, little is known about the regulation of heterologous synapsis. Here, we study the dynamics of heterologous synapsis in the nematode C. elegans. We characterize two experimental scenarios: SC assembly onto a folded-back chromosome that cannot pair with its homologous partner; and synapsis of pseudo-homologs, a fusion chromosome partnering with an unfused chromosome half its size. We observed elevated levels of heterologous synapsis when the number of meiotic double-strand breaks or crossovers were reduced, indicating that the promiscuity of synapsis is regulated by break formation or repair. In addition, our data suggests the existence of both chromosome-specific and nucleus-wide regulation on heterologous synapsis.

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Acknowledgements

We would like to thank Shawn Ahmed for worm strains; Yumi Kim and Abby Dernburg for antibodies; members of the Rog lab for discussions; Lexy Diezmann, Yuxuan Li, and Yifan Sun for statistical advice; Lisa Kursel and Yuval Mazor for critical reading of the manuscript and editorial suggestions; the Jorgensen Lab for NGM and auxin plates; the Taft-Nicholson Center for a Summer Faculty residency; and The University of Utah Cell Imaging Core for access to Imaris software.

Funding

Worm strains were provided by the CGC, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440). Work in the Rog lab is supported by R35GM128804 grant from NIGMS, and start-up funds from the University of Utah.

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HL carried out most experimental work. SGG performed the 2D analysis (Fig 3d) and made the initial observation regarding spo-11(−) animals. HL and OR analyzed the data and wrote the manuscript, with input from all authors.

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Correspondence to Ofer Rog.

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The original version of this article was revised. Missing Supplementary table and ESM legends have been added.

Supplementary information

Supplementary Fig. 1

Meiotic progression in C. elegansOverview of meiotic progression in the C. elegans gonad (dsb-2; him-8, 48 hours post-L4 gonad is shown). Dissected gonad is stained with DAPI to label DNA (blue). Meiosis progresses from left to right. The him-8 mutation, present in all strains used in this study, extends the transition zone, which spans only ~10% of the gonad in him-8(+) animals (Harper et al. 2011). The pachytene region is divided into three bins of equal length in order to observe the temporal progression of heterologous synapsis. (PNG 514 kb)

High resolution image (EPS 16605 kb)

Supplementary Fig. 2

Construction of aid::COSA-1 using CRISPR/Cas9a. Two single-stranded oligonucleotides serve as the template to create aid::COSA-1. Top: Schematic of the ultramer templates. AID degron tag (Zhang et al. 2015) is inserted at the N-terminus of COSA-1. The two templates are 150 and 125 nucleotides long (ultramers, IDT), with a 35-nucleotide overlap. The templates consist of homology to the cosa-1 5’ untranslated region, AID degron sequence, a six-amino acid linker of glycines and serines, and homology to the first exon and intron of cosa-1. The CGG PAM sequence (yellow), at the end of the first cosa-1 exon and the beginning of the first cosa-1 intron, is mutated to CGC to avoid re-cutting. Middle: the ultramer sequences. Bottom: the guide RNA sequence. Exon sequences are shown in uppercase.b. Schematic of the resulting AID::COSA-1 protein, drawn to scale. (PNG 148 kb)

High resolution image (EPS 1848 kb)

Supplementary Fig 3

Optimization and characterization of AID::COSA-1 a. gld-1 promoter is more robust than sun-1 promoter in driving AID::COSA-1 degradation. Average self-progeny is shown in the presence and absence of auxin. gld-1p::tir-1 is used as the negative control while sun-1p::tir-1 spo-11::aid him-8 (i.e. spo-11(-)) is used as a positive control. TIR-1 driven by either the sun-1 or gld-1 promoters degrades AID::COSA-1 when on auxin, yielding a lower brood size (p<0.005 compared to on NGM plates, Welch’s t-test). However, TIR-1 driven by gld-1 promotor is more robust (p=0.0475 compared with sun-1 promotor, Welch’s t-test). N represents the number of worms assessed. Genotypes are indicated at the bottom.b. Degradation of AID::COSA-1 on auxin by gld-1 promoter-driven TIR-1 yields more than six DAPI bodies in diakinesis. Numbers indicate counted DAPI bodies. Circled numbers indicate linked homologs (bivalents). Blue, DAPI bodies. Green, HTP-3 (axis). Scale bar=2 µm.c. gld-1p::tir-1; aid::COSA-1; him-8 yields similar number of DAPI bodies as zim-2 zim-3 him-8 (p=0.3976, Student’s t-test), indicating the presence of about two crossovers per meiosis (Phillips and Dernburg 2006). N represents the number of diakinesis nuclei assessed, each represented by a dot. Bars show mean ± SD. Genotypes and their effects are indicated at the bottom.(PNG 227 kb)

High resolution image (EPS 2336 kb)

Supplementary File:

Reagent ListList of all reagents used in this study, including worm strains, antibodies and primers. (XLSX 12.8 kb)

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Liu, H., Gordon, S.G. & Rog, O. Heterologous synapsis in C. elegans is regulated by meiotic double-strand breaks and crossovers. Chromosoma (2021). https://doi.org/10.1007/s00412-021-00763-y

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Keywords

  • C. elegans
  • Heterologous synapsis
  • Synaptonemal complex
  • Double-strand breaks
  • Crossover