Mapping Meiotic DNA Breaks: Two Fully-Automated Pipelines to Analyze Single-Strand DNA Sequencing Data, hotSSDS and hotSSDS-extra.

Molecular approaches are required to detect DNA double-strand break (DSB) events and to map and quantify them at high resolution. One of the most popular molecular methods in the field of meiotic recombination is the ChIP-SSDS (Chromatin immuno-precipitation and single-strand DNA sequencing). Here, we present two fully-automated Nextflow-based pipelines to analyze the sequencing data generated by this method. The first one identifies highly reproducible DSB sites, while the second provides a characterization of recovered DSB sites, including the description of the hotspot distribution and intensity along the genome and the overlap with specific regions such as gene features or known DSB hotspots. Finally, we discuss limitations/advantages and key points to consider when applying this method to specific genotypes or unconventional species.

Assessing the effects of quantitative host resistance on the life-history traits of sporulating parasites with growing lesions.

Assessing life-history traits of parasites on resistant hosts is crucial in evolutionary ecology. In the particular case of sporulating pathogens with growing lesions, phenotyping is difficult because one needs to disentangle properly pathogen spread from sporulation. By considering Phytophthora infestans on potato, we use mathematical modelling to tackle this issue and refine the assessment of pathogen response to quantitative host resistance. We elaborate a parsimonious leaf-scale model by convolving a lesion growth model and a sporulation function, after a latency period. This model is fitted to data obtained on two isolates inoculated on three cultivars with contrasted resistance level. Our results confirm a significant host-pathogen interaction on the various estimated traits, and a reduction of both pathogen spread and spore production, induced by host resistance. Most interestingly, we highlight that quantitative resistance also changes the sporulation function, the mode of which is significantly time-lagged. This alteration of the infectious period distribution on resistant hosts may have strong impacts on the dynamics of parasite populations, and should be considered when assessing the durability of disease control tactics based on plant resistance management. This inter-disciplinary work also supports the relevance of mechanistic models for analysing phenotypic data of plant-pathogen interactions.

Mouse ANKRD31 Regulates Spatiotemporal Patterning of Meiotic Recombination Initiation and Ensures Recombination between X and Y Sex Chromosomes.

Orderly segregation of chromosomes during meiosis requires that crossovers form between homologous chromosomes by recombination. Programmed DNA double-strand breaks (DSBs) initiate meiotic recombination. We identify ANKRD31 as a key component of complexes of DSB-promoting proteins that assemble on meiotic chromosome axes. Genome-wide, ANKRD31 deficiency causes delayed recombination initiation. In addition, loss of ANKRD31 alters DSB distribution because of reduced selectivity for sites that normally attract DSBs. Strikingly, ANKRD31 deficiency also abolishes uniquely high rates of recombination that normally characterize pseudoautosomal regions (PARs) of X and Y chromosomes. Consequently, sex chromosomes do not form crossovers, leading to chromosome segregation failure in ANKRD31-deficient spermatocytes. These defects co-occur with a genome-wide delay in assembling DSB-promoting proteins on autosome axes and loss of a specialized PAR-axis domain that is highly enriched for DSB-promoting proteins in wild type. Thus, we propose a model for spatiotemporal patterning of recombination by ANKRD31-dependent control of axis-associated DSB-promoting proteins.

PRDM9 Methyltransferase Activity Is Essential for Meiotic DNA Double-Strand Break Formation at Its Binding Sites.

The programmed formation of hundreds of DNA double-strand breaks (DSBs) is essential for proper meiosis and fertility. In mice and humans, the location of these breaks is determined by the meiosis-specific protein PRDM9, through the DNA-binding specificity of its zinc-finger domain. PRDM9 also has methyltransferase activity. Here, we show that this activity is required for H3K4me3 and H3K36me3 deposition and for DSB formation at PRDM9-binding sites. By analyzing mice that express two PRDM9 variants with distinct DNA-binding specificities, we show that each variant generates its own set of H3K4me3 marks independently from the other variant. Altogether, we reveal several basic principles of PRDM9-dependent DSB site determination, in which an excess of sites are designated through PRDM9 binding and subsequent histone methylation, from which a subset is selected for DSB formation.

In vivo binding of PRDM9 reveals interactions with noncanonical genomic sites.

In mouse and human meiosis, DNA double-strand breaks (DSBs) initiate homologous recombination and occur at specific sites called hotspots. The localization of these sites is determined by the sequence-specific DNA binding domain of the PRDM9 histone methyl transferase. Here, we performed an extensive analysis of PRDM9 binding in mouse spermatocytes. Unexpectedly, we identified a noncanonical recruitment of PRDM9 to sites that lack recombination activity and the PRDM9 binding consensus motif. These sites include gene promoters, where PRDM9 is recruited in a DSB-dependent manner. Another subset reveals DSB-independent interactions between PRDM9 and genomic sites, such as the binding sites for the insulator protein CTCF. We propose that these DSB-independent sites result from interactions between hotspot-bound PRDM9 and genomic sequences located on the chromosome axis.

Private selective sweeps identified from next-generation pool-sequencing reveal convergent pathways under selection in two inbred Schistosoma mansoni strains.

BACKGROUND: The trematode flatworms of the genus Schistosoma, the causative agents of schistosomiasis, are among the most prevalent parasites in humans, affecting more than 200 million people worldwide. In this study, we focused on two well-characterized strains of S. mansoni, to explore signatures of selection. Both strains are highly inbred and exhibit differences in life history traits, in particular in their compatibility with the intermediate host Biomphalaria glabrata. METHODOLOGY/PRINCIPAL FINDINGS: We performed high throughput sequencing of DNA from pools of individuals of each strain using Illumina technology and identified single nucleotide polymorphisms (SNP) and copy number variations (CNV). In total, 708,898 SNPs were identified and roughly 2,000 CNVs. The SNPs revealed low nucleotide diversity (π = 2 × 10(-4)) within each strain and a high differentiation level (Fst = 0.73) between them. Based on a recently developed in-silico approach, we further detected 12 and 19 private (i.e. specific non-overlapping) selective sweeps among the 121 and 151 sweeps found in total for each strain. CONCLUSIONS/SIGNIFICANCE: Functional annotation of transcripts lying in the private selective sweeps revealed specific selection for functions related to parasitic interaction (e.g. cell-cell adhesion or redox reactions). Despite high differentiation between strains, we identified evolutionary convergence of genes related to proteolysis, known as a key virulence factor and a potential target of drug and vaccine development. Our data show that pool-sequencing can be used for the detection of selective sweeps in parasite populations and enables one to identify biological functions under selection.

To be or not to be solitary: Phytophthora infestans' dilemma for optimizing its reproductive fitness in multiple infections.

The success of parasitic life lies in an optimal exploitation of the host to satisfy key functions directly involved in reproductive fitness. Resource availability generally decreases over time with host mortality, but also during multiple infections, where different strains of parasite share host resources. During multiple infections, the number of parasite strains and their genetic relatedness are known to influence their reproductive rates. Using infections of the potato plant Solanum tuberosum with the parasite Phytophthora infestans, we set up an experimental design to separate dose effects (double- vs. single-site infections) from genetic relatedness (different vs. identical genotypes) on the reproductive fitness of competing parasite genotypes. We showed the existence of two basic response patterns--increase or decrease in reproductive fitness in multiple infections- depending on the parasite genotype. In all cases, the intensity of the response of any genotype depended on the genotype of the competing strain. This diversity of responses to multiple infections is probably maintained by the fluctuating frequencies of multiple infections in nature, arising from variations in disease pressure over the course of an epidemic and between successive epidemics. It allows a rapid response of parasitic populations to changing environments, which are particularly intense in agricultural systems.