De novo germline mutations arise preferentially in male owing to fundamental differences between spermatogenesis and oogenesis. Post-meiotic chromatin remodeling in spermatids results in the elimination of most of the nucleosomal supercoiling and is characterized by transient DNA fragmentation. Using three alternative methods, DNA from sorted populations of mouse spermatids was used to confirm that double-strand breaks (DSB) are created in elongating spermatids and repaired at later steps. Specific capture of DSB was used for whole-genome mapping of DSB hotspots (breakome) for each population of differentiating spermatids. Hotspots are observed preferentially within introns and repeated sequences hence are more prevalent in the Y chromosome. When hotspots arise within genes, those involved in neurodevelopmental pathways become preferentially targeted reaching a high level of significance. Given the non-templated DNA repair in haploid spermatids, transient DSBs formation may, therefore, represent an important component of the male mutation bias and the etiology of neurological disorders, adding to the genetic variation provided by meiosis.

The near complete replacement of somatic chromatin in spermatids is, perhaps, the most striking nuclear event known to the eukaryotic domain. The process is far from being fully understood, but research has nevertheless unraveled its complexity as an expression of histone variants and post-translational modifications that must be finely orchestrated to promote the DNA topological change and compaction provided by the deposition of protamines. That this major transition may not be genetically inert came from early observations that transient DNA strand breaks were detected in situ at chromatin remodeling steps. The potential for genetic instability was later emphasized by our demonstration that a significant number of DNA double-strand breaks (DSBs) are formed and then repaired in the haploid context of spermatids. The detection of DNA breaks by 3′OH end labeling in the whole population of spermatids suggests that a reversible enzymatic process is involved, which differs from canonical apoptosis. We have set the stage for a better characterization of the genetic impact of this transition by showing that post-meiotic DNA fragmentation is conserved from human to yeast, and by providing tools for the initial mapping of the genome-wide DSB distribution in the mouse model. Hence, the molecular mechanism of post-meiotic DSB formation and repair in spermatids may prove to be a significant component of the well-known male mutation bias. Based on our recent observations and a survey of the literature, we propose that the chromatin remodeling in spermatids offers a proper context for the induction of de novo polymorphism and structural variations that can be transmitted to the next generation.

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