Fine-Scale Study of Meiotic Recombination in Cattle

peer reviewe

Benchmarking phasing software with a whole-genome sequenced cattle pedigree.

peer reviewed[en] BACKGROUND: Accurate haplotype reconstruction is required in many applications in quantitative and population genomics. Different phasing methods are available but their accuracy must be evaluated for samples with different properties (population structure, marker density, etc.). We herein took advantage of whole-genome sequence data available for a Holstein cattle pedigree containing 264 individuals, including 98 trios, to evaluate several population-based phasing methods. This data represents a typical example of a livestock population, with low effective population size, high levels of relatedness and long-range linkage disequilibrium. RESULTS: After stringent filtering of our sequence data, we evaluated several population-based phasing programs including one or more versions of AlphaPhase, ShapeIT, Beagle, Eagle and FImpute. To that end we used 98 individuals having both parents sequenced for validation. Their haplotypes reconstructed based on Mendelian segregation rules were considered the gold standard to assess the performance of population-based methods in two scenarios. In the first one, only these 98 individuals were phased, while in the second one, all the 264 sequenced individuals were phased simultaneously, ignoring the pedigree relationships. We assessed phasing accuracy based on switch error counts (SEC) and rates (SER), lengths of correctly phased haplotypes and the probability that there is no phasing error between a pair of SNPs as a function of their distance. For most evaluated metrics or scenarios, the best software was either ShapeIT4.1 or Beagle5.2, both methods resulting in particularly high phasing accuracies. For instance, ShapeIT4.1 achieved a median SEC of 50 per individual and a mean haplotype block length of 24.1 Mb (scenario 2). These statistics are remarkable since the methods were evaluated with a map of 8,400,000 SNPs, and this corresponds to only one switch error every 40,000 phased informative markers. When more relatives were included in the data (scenario 2), FImpute3.0 reconstructed extremely long segments without errors. CONCLUSIONS: We report extremely high phasing accuracies in a typical livestock sample. ShapeIT4.1 and Beagle5.2 proved to be the most accurate, particularly for phasing long segments and in the first scenario. Nevertheless, most tools achieved high accuracy at short distances and would be suitable for applications requiring only local haplotypes

A validation study of loci associated with mastitis resistance in two French dairy sheep breeds

The identification of loci associated with resistance to mastitis or of the causative mutations may be helpful in breeding programs for dairy sheep as it is for cattle worldwide. Seven genomic regions that control milk somatic cell counts, an indirect indicator of udder infection, have already been identified in sheep (Spanish Churra, French Lacaune and Italian Sardinian-Lacaune backcross populations). In this study, we used a 960 custom-designed ovine single nucleotide polymorphism (SNP) chip in Lacaune and Manech Tete Rousse dairy sheep to validate these seven genomic regions associated with mastitis. The most significant SNP (rs868996547) on Ovis aries chromosome (OAR) 3 was a previously described mutation in the suppressor of cytokine signalling 2 (SOCS2) gene. An antagonist effect of this causal candidate between health and growth in Lacaune sheep was confirmed. Effects of the mutation on the infectious status of the udder, i.e. increases in milk somatic cell counts and bacteria shedding, were also identified. This SNP was not present in the data available on Manech Tete Rousse. Three other regions associated with mastitis were also confirmed on OAR16 (Manech Tete Rousse), 19 (Lacaune) and 2 (both breeds). For the OAR2 region, we validated previously detected SNPs in several other breeds (Sarda, Churra, and Chios). For significant SNPs in the four mastitis regions, the effect varied from 0.24 to 0.67 phenotypic standard deviation of the traits. Two of the mastitis quantitative trait loci (QTL) regions (OAR2 and 16) that we validated here were also associated in opposite ways with milk production traits in both populations. These results indicate, at least in part, a genomic basis for the trade-off between milk production and mastitis resistance. Four of the seven mastitis QTL regions that were previously identified in independent populations, were confirmed in this study, which demonstrates partial sharing of mastitis-related genetic mechanisms between different distant dairy sheep populations

Rare CNVs in the bovine genome are not captured well by 50K density genotyping array SNPs

peer reviewe

High-resolution structural variants catalogue in a large-scale whole genome sequenced bovine family cohort data.

peer reviewed[en] BACKGROUND: Structural variants (SVs) are chromosomal segments that differ between genomes, such as deletions, duplications, insertions, inversions and translocations. The genomics revolution enabled the discovery of sub-microscopic SVs via array and whole-genome sequencing (WGS) data, paving the way to unravel the functional impact of SVs. Recent human expression QTL mapping studies demonstrated that SVs play a disproportionally large role in altering gene expression, underlining the importance of including SVs in genetic analyses. Therefore, this study aimed to generate and explore a high-quality bovine SV catalogue exploiting a unique cattle family cohort data (total 266 samples, forming 127 trios). RESULTS: We curated 13,731 SVs segregating in the population, consisting of 12,201 deletions, 1,509 duplications, and 21 multi-allelic CNVs (> 50-bp). Of these, we validated a subset of copy number variants (CNVs) utilising a direct genotyping approach in an independent cohort, indicating that at least 62% of the CNVs are true variants, segregating in the population. Among gene-disrupting SVs, we prioritised two likely high impact duplications, encompassing ORM1 and POPDC3 genes, respectively. Liver expression QTL mapping results revealed that these duplications are likely causing altered gene expression, confirming the functional importance of SVs. Although most of the accurately genotyped CNVs are tagged by single nucleotide polymorphisms (SNPs) ascertained in WGS data, most CNVs were not captured by individual SNPs obtained from a 50K genotyping array. CONCLUSION: We generated a high-quality SV catalogue exploiting unique whole genome sequenced bovine family cohort data. Two high impact duplications upregulating the ORM1 and POPDC3 are putative candidates for postpartum feed intake and hoof health traits, thus warranting further investigation. Generally, CNVs were in low LD with SNPs on the 50K array. Hence, it remains crucial to incorporate CNVs via means other than tagging SNPs, such as investigation of tagging haplotypes, direct imputation of CNVs, or direct genotyping as done in the current study. The SV catalogue and the custom genotyping array generated in the current study will serve as valuable resources accelerating utilisation of full spectrum of genetic variants in bovine genomes.Seventh Framework ProgrammeH202

Enrichment of causative variants in tissue-specific and shared ATAC-Seq peaks in cattle

peer reviewe

Genetic diversity analysis of French goat populations reveals soft selective sweeps involved in their adaptation

Supplementary data for publication: Oget C., Servin B., Palhiere I. (2019). Genetic diversity analysis of French goat populations reveals soft selective sweeps involved in their adaptation. Animal Genetics, Volume 50, Issue 1

Effet pléiotrope de la mutation R96C dans le gène SOCS2 chez la brebis laitière

In this PhD thesis, we investigated a mutation in the SOCS2gene (Suppressor Of Cytokine Signaling 2) with a pleiotropic effect, i.e.it affects several traits of interest in the Lacaune dairy sheep. This mutation is unfavorable for mastitis resistance (udderinflammation), and favorable for growth and milk production traits.This point mutation (SNP -Single Nucleotide Polymorphism), located in the highly conserved binding domain of the SOCS-2 protein,causes the loss of functionality of the protein that is involved in a major signaling pathway in mammals:the JAK(Janus Kinase) / STAT (Signal Transducer and Activator of Transcription) pathway, regulating a large spectrum of cytokines and growth factors.The first objective of this PhD thesis was to provide a basis for considering this mutation in the context of genomic selection in the Lacaune breed. Weighted evaluation methods, and the addition of the SOCS2SNP among the chip markers, brought gains in accuracy on predictions (+3.99% and +0.26% respectively on average), suggesting the possibility of more efficient selection. The Gene Contentmethod was also interesting because it alloweddissociatingthe genetic value due to the SOCS2gene from that of the other genes (polygene). Thus, we have shown that the current selection in Lacaune breed allows reducing the frequency of the unfavorable SOCS2allele while improving the resistance to mastitis explained by the remaining part of the genome.The second objective was to study the effects of SOCS2on traits of interest not yet investigated: udder infection status and reproduction in females, and growth in males. Association studies with a 960 SNP chip, including SOCS2SNP, confirmed the effect of SOCS2on milk production and mammary inflammation, and showed a direct effect on infection using new fine phenotypes (bacteriology, clinical examinations). Linear models showed that the SOCS2mutation was associated withan increase in weight in young males (+1.5%), as observed in ewes, and a decrease in the artificial insemination success rate in ewes (+1.3 times the risk of failure), with no effect on prolificacy. These striking and original results on reproduction pave the way for further analyses to determine in particular at what biological stage (ovulation, fecundation, embryonic implantation, etc.) the SOCS-2 protein could be involved.The last objective was to determine the role of the SOCS-2 protein in the underlying biological mechanisms of immunity by investigating the response of 14 homozygous carrier and non-carrier ewes to intramammary infection by Staphylococcus aureus. All ewes developed two peaks of immune cell recruitment to the udder at 20 and 88 hours post-inoculation (T20 and T88). Differential analysis of the expressed genes confirmed a modification of the transcriptome from T16 related to the activation of the immune system, the DNA repair and the cellular apoptosis pathways. Ewes carrying the SOCS2mutation were distinguished from wild ewes by an increased clinical response at T56 and a difference in T lymphocyte recruitment at the time of the inflammatory peaks. At T56, a total of 177 genes were differentially expressed between the two genotypes. A functional analysis of these genes, and the integration between gene expression and phenotypes, suggest that the mutation leads to an over activation of signal transmission and regulatory pathways (interferons, STAT3) with a deleterious impact on the clinical status of ewe.Dans cette thèse, nous nous sommes intéressée à une mutation dans le gène SOCS2(Suppressor Of Cytokine Signaling 2) dont l’effet est pléiotrope, c’est-à-dire qu’il concerne plusieurs caractères d’intérêt chez la brebis laitière Lacaune. Cette mutation estdéfavorable pour la résistance aux mammites (inflammation de la mamelle), et favorable pour les caractères de croissance et de productionlaitière. Située dans le domaine de liaison très conservé de la protéine SOCS-2, cette mutation ponctuelle (SNP:Single Nucleotide Polymorphism) entraîne la perte de fonctionnalité de la protéine qui est impliquée dans une voie majeure de signalisation chez les mammifères, la voie JAK (Janus Kinase) / STAT (Signal Transducer and Activator of Transcription), régulant un large spectre de cytokines et de facteurs de croissance.Le premier objectif de cette thèse était de proposer la prise en compte de cette mutation dans le contexte de la sélection génomique dans la race Lacaune. Les méthodes d’évaluation pondérées, et l’ajout du SNP SOCS2parmi les marqueurs de la puce, ont apporté des gains de précisions sur les prédictions (respectivement +3,99% et +0,26% en moyenne), laissant envisager la possibilité d’unesélection plus efficace. La méthode Gene Contentétait également intéressante car elle a permis de dissocier la valeur génétique due au gène SOCS2de celle provenant des autres gènes (polygène). Ainsi nous avons montré que la sélection actuelle en race Lacaune permet de diminuer la fréquence de l’allèle défavorable SOCS2tout en améliorant la résistance aux mammites expliquée par le reste du génome.Le deuxième objectif était d’étudier les effets de SOCS-2 sur des caractères d’intérêt non évalués jusqu’à présent: statut infectieux des mamelles et reproduction chez la femelle, et croissance des mâles. Des études d’association avec une puce de 960 SNP, dont le SNP SOCS2, ont confirmé l’effet de SOCS2sur la production laitière et sur l’inflammation mammaire, et ont montré un effet direct sur l’infection à l’aide de nouveaux phénotypes fins (bactériologie, examens cliniques). Des modèles linéaires ont montréque la mutation SOCS2était associée à une augmentation de poids chez les jeunes mâles (+1,5%), comme cela avait été observé chez la brebis, et àune diminution du taux de réussite à l’insémination artificielle chez les brebis (+1,3 fois de risque d’échec), sans effet sur la prolificité. Ces résultats marquants et originaux sur la reproduction ouvrent la voie à des analyses complémentaires pour déterminer notamment à quelstade biologique (ovulation, fécondation, implantation embryonnaire, etc.) la protéine SOCS-2 pourrait intervenir.Le dernier objectif était de déterminer le rôle de la protéine SOCS-2 dans les mécanismes biologiques sous-jacents de l’immunité grâce à l’étude de la réponse de 14 brebis porteuses et non porteuses de la mutation à l’état homozygote dans le cadre d’une infection intra-mammaire par un Staphylococcus aureus. L’ensemble des brebis a présenté deux pics de recrutement de cellules immunitaires versla mamelle à 20 et 88 heures post-inoculation (T20 et T88). Une analyse différentielle des gènes exprimés a confirmé une modification du transcriptome à partir de T16 en lien avec l’activation du système immunitaire,des voies de réparation de l’ADN et d’apoptose cellulaire. Les brebis portant la mutation SOCS2se sont différenciées des brebis sauvages, par une réaction clinique accrue à T56 et une différence de recrutement de lymphocytes T au moment des pics inflammatoires. A T56, un total de 177 gènes était différentiellement exprimé entre les deux génotypes. Une analyse fonctionnelle de ces gènes, et l’intégration entre l’expression génique et les phénotypes, suggèrent que la mutation conduit à une sur activation des voies de transmission de signal et derégulation (interférons, STAT3) avec un impact délétère sur l’état clinique des brebis

Pleiotropic effect of R96C SOCS2 mutation in dairy sheep

Les mammites représentent un gros fardeau pour l'industrie laitière en raison de l'altération de la qualité du lait et de l'augmentation du coût de production. En plus des mesures d'hygiène mises en oeuvre, la sélection en faveur d'une plus grande immunité contre les infections mammaires et de la résistance aux mammites est considérée et implémentée dans les programmes d'amélioration de plusieurs races laitières. Cependant,la résistance aux mammites est un caractère complexe dont les bases biologiques de détermination génétique restent peu connues. Depuis une dizaine d'années, le décryptage des bases génétiques et immunologiques des mammites est un sujet de recherche majeur pour les unités partenaires GenPhySE et IHAP de l'INRA. A partir d'études d'association pangénomique sur le taux de cellules somatiques dans le lait, un caractère lié aux mammites, ils ont identifié une mutation ponctuelle dans le domaine de liaison SH2 du gène SOCS-2, entraînant la perte de reconnaissance du ligand (Rupp et al., 2015). La fréquence de cette mutation était étonnamment élevée (21%) dans la population étudiée. De plus, la taille, le poids et la production laitière étaient significativement augmentés chez les brebis porteuses de l'allèle mutant socs2 en comparaison au phénotype sauvage. Tous ces résultats ont apporté des preuves solides en faveur d'une mutation causale contrôlant l'inflammation mammaire chez le mouton et ont mis en lumière le rôle majeur du gène SOCS-2 comme étant un potentiel facteur concurrentiel entre la réponse inflammatoire provoquée par des pathogènes lors des infections mammaires et la production laitière. Le projet de thèse s'inscrit dans le cadre des travaux en cours sur l'étude du rôle du gène SOCS-2 dans la prédisposition aux infections bactériennes, et de ses effets pléiotropes à la fois sur la santé et sur les caractères de production, au sein du projet ANR REIDSOCS (2016-2020). Le premier objectif est d'établir la fréquence de l'allèle "T" muté socs2 associé à une augmentation de la prédisposition aux mammites dans la population ovine. Les relations entre mammites et caractères de production (lait et croissance) seront étudiées afin d'émettre des hypothèses concernant les forces de sélection qui ont conduit à la propagation de l'allèle 'T' délétère socs2 dans la population ovine. Des analyses d'association pangénomique sur de nouveaux phénotypes (bactériologie du lait) permettront ensuite d'évaluer les effets du gène SOCS-2 et d'autres régions du génome sur le caractère de résistance aux mammites. Puis, le deuxième objectif est de réaliser une étude prospective sur la gestion de cette mutation. Diverses méthodes seront testées en conséquence afin de représenter la mutation au sein de l'évaluation génomique actuelle et à pour l'optimisation du schéma de sélection. • Le troisième objectif est d'explorer les gènes et mécanismes sous-jacents de la réponse aux infections modifiée en lien avec la mutation ponctuelle du gène SOCS-2. Des génotypes d'individus des variants Socs2 seront produits au sein d'une ferme expérimentale INRA afin de réaliser des profils de transcription après des tests expérimentaux. Enfin, un dépistage systématique du polymorphisme de Socs2 chez les espèces de ruminants (bovins, ovins, caprins) sera effectué à l'aide d'analyses in silico de données de séquence publiques. La dernière étude sera étendue à des gènes en interaction avec le gène SOCS-2 qui seront identifiés par d'autres partenaires du projet REIDSOCS.In this PhD thesis, we investigated a mutation in the SOCS2 gene (Suppressor Of Cytokine Signaling 2) with a pleiotropic effect, i.e. it affects several traits of interest in the Lacaune dairy sheep. This mutation is unfavourable for mastitis resistance (udder inflammation), and favourable for growth and milk production traits..T Located in the highly conserved binding domain of the SOCS2 protein, this point mutation (SNP - Single Nucleotide Polymorphism), l ocated in the highly conserved binding domain of the SOCS2 protein, causes the loss of functionality of the protein that is involved in a major signalling pathway in mammals:, the JAK (Janus Kinase) / STAT (Signal Transducer and Activator of Transcription) pathway, regulating a large spectrum of cytokines and growth factors. The first objective of this PhD thesis was to provide a basis for considering this mutation in the context of genomic selection in the Lacaune breed. Weighted evaluation methods, and the addition of the SOCS2 SNP among the chip markers, brought gains in accuracy on predictions (+3.99% and +0.26% respectively on average), suggesting the possibility of more efficient selection. The Gene Content method was also interesting because it made it possibleallowed to dissociate the genetic value due to the SOCS2 gene from that of the other genes (polygene). Thus, we have shown that the current selection in Lacaune breed allows reducing the frequency of the unfavourable SOCS2 allele while improving the resistance to mastitis explained by the remaining part of the genome. The second objective was to study the effects of SOCS2 on traits of interest not yet investigated: udder infection status and reproduction in females, and growth in males. Association studies with a 960 SNP chip, including SOCS2 SNP, confirmed the effect of SOCS2 on milk production and mammary inflammation, and showed a direct effect on infection using new fine phenotypes (bacteriology, clinical examinations). Linear models showed that the SOCS2 mutation was associated with: i) an increase in weight in young males (+1.5%), as observed in ewes, and ii) a decrease in the artificial insemination success rate in ewes (+1.3 times the risk of failure), with no effect on prolificacy. These striking and original results on reproduction pave the way for further analyses to determine in particular at what biological stage (ovulation, fecundation, embryonic implantation, etc.) the SOCS2 protein could be involved. The last objective was to determine the role of the SOCS2 protein in the underlying biological immune mechanisms of immunity by investigating the response of 14 homozygous carrier and non-carrier ewes to intramammary infection by Staphylococcus aureus. All ewes developed two peaks of immune cell recruitment to the udder at 20 and 88 hours post-inoculation (T20 and T88). Differential analysis of the expressed genes confirmed a modification of the transcriptome from T16 related to the activation of the immune system, the DNA repair and the cellular apoptosis pathways. Ewes carrying the SOCS2 mutation were distinguished from wild ewes by an increased clinical response at T56 and a difference in T lymphocyte recruitment at the time of the inflammatory peaks. At T56, a total of 177 genes were differentially expressed between the two genotypes. A functional analysis of these genes, and the integration between gene expression and phenotypes, suggest that the mutation leads to an over activation of signal transmission and regulatory pathways (interferons, STAT3) with a deleterious impact on the clinical status of ewes