A simple and rapid method for imaging male meiotic cells in anthers of model and non-model plant species

Roßig, Claudia; Lievre, Liam Le; Pilkington, Sarah M.; Brownfield, Lynette

doi:10.1007/s00497-021-00404-5

Cited by 6 publications

(6 citation statements)

References 30 publications

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“…2C, colored dots), which was confirmed by a significant anther effect (mixed-effects model, P<0.001). These similar average speeds between centromeres of the same anther revealed some degree of synchronization in the average speed between meiocytes, which could be a direct consequence of the synchronization of meiosis within an anther 43,44 . We then asked whether such coordination could occur between the centromeres of a given meiocyte.…”

Section: A Thaliana Centromere Movements Are Poorly Coordinated Withi...mentioning

confidence: 75%

Rapid meiotic prophase chromosome movements inArabidopsis thalianaare linked to essential reorganization at the nuclear envelope

Cromer,

Tiscareno-Andrade,

Lefranc

et al. 2023

Preprint

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Meiotic rapid prophase chromosome movements (RPMs) require connections between the chromosomes and the cytoskeleton, involving SUN (Sad1/UNC-84)-domain-containing proteins at the inner nuclear envelope (NE). RPMs remain significantly understudied in plants, notably with respect to their importance in the regulation of meiosis. Here, we demonstrate thatArabidopsis thalianameiotic centromeres undergo rapid (up to 500 nm/s) and uncoordinated movements during the zygotene and pachytene stages. RPMs are not affected by altered chromosome organization and recombination but are abolished in the double mutantsun1 sun2. We also document the changes in chromosome dynamics and nucleus organization during the transition from leptotene to zygotene, including telomere attachment to SUN-enriched NE domains, bouquet formation, and nucleolus displacement, all of which were defective insun1 sun2. These results establishA. thalianaas a new model species for studying the functional implications of meiotic RPMs and demonstrate the mechanistic conservation of telomere-led RPMs in plants.

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Section: A Thaliana Centromere Movements Are Poorly Coordinated Withi...mentioning

confidence: 75%

Rapid meiotic prophase chromosome movements inArabidopsis thalianaare linked to essential reorganization at the nuclear envelope

Cromer,

Tiscareno-Andrade,

Lefranc

et al. 2023

Preprint

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“…In Arabidopsis, while each inflorescence contains flower buds bearing PMC at different pre-meiotic and meiotic stages, the PMC of the anthers of a given flower bud are nearly synchronous [25,26]. Along with the sequence of cytologically distinguishable meiotic stages in the buds of a given inflorescence, in practice it is relatively straightforward to classify PMC subpopulations within the meiotic timeline.…”

Section: Resultsmentioning

confidence: 99%

“…The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. 26…”

Section: Acknowledgementsmentioning

confidence: 99%

Meiotic DSB repair DNA synthesis tracts inArabidopsis thaliana

Hernández Sánchez-Rebato,

Schubert,

White

2024

Preprint

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We report here the successful labelling of meiotic prophase I DNA synthesis in the flowering plant,Arabidopsis thaliana. Incorporation of the thymidine analogue, EdU, enables visualisation of the footprints of recombinational repair of programmed meiotic DNA double-strand breaks (DSB), with ∼400 discrete, SPO11-dependent, EdU-labelled chromosomal foci clearly visible at pachytene and later stages of meiosis. This number equates well with previous estimations of 200-300 DNA double-strand breaks per meiosis in Arabidopsis, confirming the power of this approach to detect the repair of most or all SPO11-dependent meiotic DSB repair recombination. The chromosomal distribution of these DNA-synthesis foci accords with that of early recombination markers and MLH1, which marks Class I crossover sites, colocalises with the EdU foci. ArabidopsisIt is currently estimated that ∼10 cross-overs (CO) and an equivalent number of non-cross-overs (NCO) occur in each Arabidopsis male meiosis. Thus, at least 90% of meiotic recombination events, and very probably more, have not previously been accessible for analysis. Visual examination of the patterns of the foci on the synapsed pachytene chromosomes corresponds well with expectations from the different mechanisms of meiotic recombination and notably, no evidence for long Break-Induced Replication DNA synthesis tracts was found. Labelling of meiotic prophase I, SPO11-dependent DNA synthesis holds great promise for further understanding of the molecular mechanisms of meiotic recombination, at the heart of reproduction and evolution of eukaryotes.Author SummarySexual reproduction involves the fusion of two cells, one from each parent. To maintain a stable chromosome complement across generations, these specialized reproductive cells must be produced through a specialized cell division called meiosis. Meiosis halves the chromosome complement of gametes and recombines the parental genetic contributions in each gamete, generating the genetic variation that drives evolution. The complex mechanisms of meiotic recombination have been intensely studied for many years and we now know that it involves the repair of programmed chromosomal breaks through recombination with intact template DNA sequences on another chromatid. At the molecular level, this is known to involve new DNA synthesis at the sites of repair/recombination and we report here the successful identification and characterisation of this DNA neo-synthesis during meiosis in the flowering plant, Arabidopsis. Both the characteristics and numbers of these DNA synthesis tracts accord with expectations from theory and earlier studies. Potentially applicable to studies in many organisms, this approach provides indelible footprints in the chromosomes and has the great advantage of freeing researchers from dependence on indirect methods involving detection of proteins involved in these dynamic processes.

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“…The analysis of meiocytes located in the anther was performed according to the published protocol (Rossig et al ., 2021). Briefly, inflorescences with closed buds were fixed overnight in 3 : 1, ethanol : acetic acid.…”

Section: Methodsmentioning

confidence: 99%

“…While multiple anthers are on each slide, the stages of interest (metaphase II) are of short duration. Slides were screened for anther locules containing this stage (locules are not all synchronised, Rossig et al ., 2021). Within anther locules containing male meiocytes at metaphase II, only meiocytes with two clear groups of chromosomes were chosen for imaging and measurement.…”

Section: Methodsmentioning

confidence: 99%

Meiocyte size is a determining factor for unreduced gamete formation in Arabidopsis thaliana

Kradolfer

Brownfield

et al. 2022

New Phytologist

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Polyploidy, the presence of more than two sets of chromosomes within a cell, is a widespread phenomenon in plants. The main route to polyploidy is considered through the production of unreduced gametes that are formed as a consequence of meiotic defects. Nevertheless, for reasons poorly understood, the frequency of unreduced gamete formation differs substantially among different plant species. The previously identified meiotic mutant jason (jas) in Arabidopsis thaliana forms about 60% diploid (2n) pollen. JAS is required to maintain an organelle band as a physical barrier between the two meiotic spindles, preventing previously separated chromosome groups from uniting into a single cell.In this study, we characterized the jas suppressor mutant telamon (tel) that restored the production of haploid pollen in the jas background.The tel mutant did not restore the organelle band, but enlarged the size of male jas tel meiocytes, suggesting that enlarged meiocytes can bypass the requirement of the organelle band. Consistently, enlarged meiocytes generated by a tetraploid jas mutant formed reduced gametes.The results reveal that meiocyte size impacts chromosome segregation in meiosis II, suggesting an alternative way to maintain the ploidy stability in meiosis during evolution.

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A simple and rapid method for imaging male meiotic cells in anthers of model and non-model plant species

Cited by 6 publications

References 30 publications

Rapid meiotic prophase chromosome movements inArabidopsis thalianaare linked to essential reorganization at the nuclear envelope

Rapid meiotic prophase chromosome movements inArabidopsis thalianaare linked to essential reorganization at the nuclear envelope

Meiotic DSB repair DNA synthesis tracts inArabidopsis thaliana

Meiocyte size is a determining factor for unreduced gamete formation in Arabidopsis thaliana

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