CENP-A binding domains and recombination patterns in horse spermatocytes

Abstract

Centromeres exert an inhibitory effect on meiotic recombination, but the possible contribution of satellite DNA to this “centromere effect” is under debate. In the horse, satellite DNA is present at all centromeres with the exception of the one from chromosome 11. This organization of centromeres allowed us to investigate the role of satellite DNA on recombination suppression in horse spermatocytes at the stage of pachytene. To this aim we analysed the distribution of the MLH1 protein, marker of recombination foci, relative to CENP-A, marker of centromeric function. We demonstrated that the satellite-less centromere of chromosome 11 causes crossover suppression, similarly to satellite-based centromeres. These results suggest that the centromere effect does not depend on satellite DNA. During this analysis, we observed a peculiar phenomenon: while, as expected, the centromere of the majority of meiotic bivalent chromosomes was labelled with a single immunofluorescence centromeric signal, double-spotted or extended signals were also detected. Their number varied from 0 to 7 in different cells. This observation can be explained by positional variation of the centromeric domain on the two homologs and/or misalignment of pericentromeric satellite DNA arrays during homolog pairing confirming the great plasticity of equine centromeres.

The impact of chromosomal fusions on 3D genome folding and recombination in the germ line

“…Although this was consistent with our observations (the proportion of chromosome arms with zero CO was slightly higher in heterozygous Rb fusions than nonfused chromosomes), we also detected a strong reduction of recombination in homozygous Rb-fused chromosomes, mirroring early cytological observations 38 . Decreased recombination in homozygous Rb fusions is counterintuitive but could be explained by the presence of double-centromeric signals that represented misaligned centromeres 39 . Since centromeres can reduce recombination rates (so-called centromere interference 40 ), the presence of double centromeres could magnify this effect, interfering with both the formation of COs during prophase and subsequently affecting chromosome segregation in metaphase I.…”

Neocentromere formation through Robertsonian fusion and centromere repositioning during the evolution of zebras

“…Chromatin from primary fibroblasts was cross-linked with 1% formaldehyde, extracted, and sonicated to obtain DNA fragments ranging from 200 to 800 bp. Immunoprecipitation was performed as previously described (Nergadze, et al 2018) by using an anti-CENP-A serum (Cappelletti, et al 2019). Grevy's zebras was performed using an iterative chromosome walking approach based on paired-end ChIPseq reads, as previously described (Nergadze, et al 2018).…”

“…Chromatin from primary fibroblasts was cross-linked with 1% formaldehyde, extracted, and sonicated to obtain DNA fragments ranging from 200 to 800 bp. Immunoprecipitation was performed as previously described (Nergadze, et al 2018) by using an anti-CENP-A serum (Cappelletti, et al 2019).…”

Decoding the Equine Genome: Lessons from ENCODE

“…The position of the ECA11 satellite-less centromere, identified as the CENP-A binding domain, is not fixed in the horse population but slides within an about the 500 kb region, giving rise to different positional alleles or "epialleles" [106,111,112]. The analysis of these epialleles carried out on families composed by horses, donkeys, and their hybrid offspring (mule/hinny) revealed that they are inherited as Mendelian traits, but their position can slide in one generation [106].…”

“…The absence of satellite DNA at these centromeres also provides a unique opportunity to understand whether some typical features of mammalian centromeres depend on the presence of satellite DNA. In particular, it was possible to demonstrate that satellite DNA was not necessary for segregation fidelity of the centromere [113] and was not implicated in the suppression of meiotic recombination, which is typically exerted by the centromere [112].…”