Pyrimidine de novo synthesis inhibition selectively blocks effector but not memory T cell development.

Blocking pyrimidine de novo synthesis by inhibiting dihydroorotate dehydrogenase is used to treat autoimmunity and prevent expansion of rapidly dividing cell populations including activated T cells. Here we show memory T cell precursors are resistant to pyrimidine starvation. Although the treatment effectively blocked effector T cells, the number, function and transcriptional profile of memory T cells and their precursors were unaffected. This effect occurred in a narrow time window in the early T cell expansion phase when developing effector, but not memory precursor, T cells are vulnerable to pyrimidine starvation. This vulnerability stems from a higher proliferative rate of early effector T cells as well as lower pyrimidine synthesis capacity when compared with memory precursors. This differential sensitivity is a drug-targetable checkpoint that efficiently diminishes effector T cells without affecting the memory compartment. This cell fate checkpoint might therefore lead to new methods to safely manipulate effector T cell responses.

Barley shows reduced Fusarium head blight under drought and modular expression of differentially expressed genes under combined stress.

Plants often face simultaneous abiotic and biotic stress conditions; however, physiological and transcriptional responses under such combined stress conditions are still not fully understood. Spring barley (Hordeum vulgare) is susceptible to Fusarium head blight (FHB), which is strongly affected by weather conditions. We therefore studied the potential influence of drought on FHB severity and plant responses in three varieties of different susceptibility. We found strongly reduced FHB severity in susceptible varieties under drought. The number of differentially expressed genes (DEGs) and strength of transcriptomic regulation reflected the concentrations of physiological stress markers such as abscisic acid or fungal DNA contents. Infection-related gene expression was associated with susceptibility rather than resistance. Weighted gene co-expression network analysis revealed 18 modules of co-expressed genes that reflected the pathogen- or drought-response in the three varieties. A generally infection-related module contained co-expressed genes for defence, programmed cell death, and mycotoxin detoxification, indicating that the diverse genotypes used a similar defence strategy towards FHB, albeit with different degrees of success. Further, DEGs showed co-expression in drought- or genotype-associated modules that correlated with measured phytohormones or the osmolyte proline. The combination of drought stress with infection led to the highest numbers of DEGs and resulted in a modular composition of the single-stress responses rather than a specific transcriptional output.

Fibroblast growth factor induced Ucp1 expression in preadipocytes requires PGE2 biosynthesis and glycolytic flux.

High uncoupling protein 1 (Ucp1) expression is a characteristic of differentiated brown adipocytes and is linked to adipogenic differentiation. Paracrine fibroblast growth factor 8b (FGF8b) strongly induces Ucp1 transcription in white adipocytes independent of adipogenesis. Here, we report that FGF8b and other paracrine FGFs act on brown and white preadipocytes to upregulate Ucp1 expression via a FGFR1-MEK1/2-ERK1/2 axis, independent of adipogenesis. Transcriptomic analysis revealed an upregulation of prostaglandin biosynthesis and glycolysis upon Fgf8b treatment of preadipocytes. Oxylipin measurement by LC-MS/MS in FGF8b conditioned media identified prostaglandin E2 as a putative mediator of FGF8b induced Ucp1 transcription. RNA interference and pharmacological inhibition of the prostaglandin E2 biosynthetic pathway confirmed that PGE2 is causally involved in the control over Ucp1 transcription. Importantly, impairment of or failure to induce glycolytic flux blunted the induction of Ucp1, even in the presence of PGE2 . Lastly, a screening of transcription factors identified Nrf1 and Hes1 as required regulators of FGF8b induced Ucp1 expression. Thus, we conclude that paracrine FGFs co-regulate prostaglandin and glucose metabolism to induce Ucp1 expression in a Nrf1/Hes1-dependent manner in preadipocytes, revealing a novel regulatory network in control of Ucp1 expression in a formerly unrecognized cell type.

Proliferation-competent Tcf1+ CD8 T cells in dysfunctional populations are CD4 T cell help independent.

T cell maintenance in chronic infection and cancer follows a hierarchical order. Short-lived effector CD8 T cells are constitutively replaced from a proliferation-competent Tcf1-expressing progenitor population. This occurs spontaneously at low levels and increases in magnitude upon blocking PD-1 signaling. We explore how CD4 T cell help controls transition and survival of the progenitors and their progeny by utilizing single-cell RNA sequencing. Unexpectedly, absence of CD4 help caused reductions in cell numbers only among terminally differentiated cells while proliferation-competent progenitor cells remained unaffected with regard to their numbers and their overall phenotype. In fact, upon restoration of a functional CD4 compartment, the progenitors began to regenerate the effector CD8 T cells. Thus, unlike memory T cells for which secondary expansion requires CD4 T cell help, this is not a necessity for proliferation-competent progenitor cells in dysfunctional populations. Our data therefore reveals that proliferation-competent cells in dysfunctional populations show a previously unrecognized uncoupling of CD4 T cell help that is otherwise required by conventional memory T cells.

Genetics of adaptation in modern chicken.

We carried out whole genome resequencing of 127 chicken including red jungle fowl and multiple populations of commercial broilers and layers to perform a systematic screening of adaptive changes in modern chicken (Gallus gallus domesticus). We uncovered >21 million high quality SNPs of which 34% are newly detected variants. This panel comprises >115,000 predicted amino-acid altering substitutions as well as 1,100 SNPs predicted to be stop-gain or -loss, several of which reach high frequencies. Signatures of selection were investigated both through analyses of fixation and differentiation to reveal selective sweeps that may have had prominent roles during domestication and breed development. Contrasting wild and domestic chicken we confirmed selection at the BCO2 and TSHR loci and identified 34 putative sweeps co-localized with ALX1, KITLG, EPGR, IGF1, DLK1, JPT2, CRAMP1, and GLI3, among others. Analysis of enrichment between groups of wild vs. commercials and broilers vs. layers revealed a further panel of candidate genes including CORIN, SKIV2L2 implicated in pigmentation and LEPR, MEGF10 and SPEF2, suggestive of production-oriented selection. SNPs with marked allele frequency differences between wild and domestic chicken showed a highly significant deficiency in the proportion of amino-acid altering mutations (P<2.5×10-6). The results contribute to the understanding of major genetic changes that took place during the evolution of modern chickens and in poultry breeding.

A nonsense mutation of bone morphogenetic protein-15 (BMP15) causes both infertility and increased litter size in pigs.

BACKGROUND: Atypical external genitalia are often a sign of reproductive organ pathologies and infertility with both environmental or genetic causes, including karyotypic abnormalities. Genome-wide association studies (GWAS) provide a means for identifying chromosomal regions harboring deleterious DNA-variants causing such phenotypes. We performed a GWAS to unravel the causes of incidental cases of atypically small vulvae in German Landrace gilts. RESULTS: A case-control GWAS involving Illumina porcine SNP60 BeadChip-called genotypes of 17 gilts with atypically small vulvae and 1818 control animals (fertile German Landrace sows) identified a significantly associated region on the X-chromosome (P = 8.81 × 10- 43). Inspection of whole-genome sequencing data in the critical area allowed us to pinpoint a likely causal variant in the form of a nonsense mutation of bone morphogenetic protein-15 (BMP15; Sscrofa11.1_X:g.44618787C>T, BMP15:p.R212X). The mutant allele occurs at a frequency of 6.2% in the German Landrace breeding population. Homozygous gilts exhibit underdeveloped, most likely not functional ovaries and are not fertile. Male carriers do not seem to manifest defects. Heterozygous sows produce 0.41±0.02 (P=4.5 × 10-83) piglets more than wildtype animals. However, the mutant allele's positive effect on litter size accompanies a negative impact on lean meat growth. CONCLUSION: Our results provide an example for the power of GWAS in identifying the genetic causes of a fuzzy phenotype and add to the list of natural deleterious BMP15 mutations that affect fertility in a dosage-dependent manner, the first time in a poly-ovulatory species. We advise eradicating the mutant allele from the German Landrace breeding population since the adverse effects on the lean meat growth outweigh the larger litter size in heterozygous sows.

Candidate genes and gene markers for the resistance to porcine pleuropneumonia.

Actinobacillus (A.) pleuropneumoniae is one of the most important respiratory pathogens in global pig production. Antimicrobial treatment and vaccination provide only limited protection, but genetic disease resistance is a very promising alternative for sustainable prophylaxis. Previous studies have discovered multiple QTL that may explain up to 30% of phenotypic variance. Based on these findings, the aim of the present study was to use genomic sequencing to identify genetic markers for resistance to pleuropneumonia in a segregating commercial German Landrace line. 163 pigs were infected with A. pleuropneumoniae Serotype 7 through a standardized aerosol infection method. Phenotypes were accurately defined on a clinical, pathological and microbiological basis. The 58 pigs with the most extreme phenotypes were genotyped by sequencing (next-generation sequencing). SNPs were used in a genome-wide association study. The study identified genome-wide associated SNPs on three chromosomes, two of which were chromosomes of QTL which had been mapped in a recent experiment. Each variant explained up to 20% of the total phenotypic variance. Combined, the three variants explained 52.8% of the variance. The SNPs are located in genes involved in the pathomechanism of pleuropneumonia. This study confirms the genetic background for the host's resistance to pleuropneumonia and indicates a potential role of three candidates on SSC2, SSC12 and SSC15. Favorable gene variants are segregating in commercial populations. Further work is needed to verify the results in a controlled study and to identify the functional QTN.

DEUS: an R package for accurate small RNA profiling based on differential expression of unique sequences.

SUMMARY: Despite their fundamental role in various biological processes, the analysis of small RNA sequencing data remains a challenging task. Major obstacles arise when short RNA sequences map to multiple locations in the genome, align to regions that are not annotated or underwent post-transcriptional changes which hamper accurate mapping. In order to tackle these issues, we present a novel profiling strategy that circumvents the need for read mapping to a reference genome by utilizing the actual read sequences to determine expression intensities. After differential expression analysis of individual sequence counts, significant sequences are annotated against user defined feature databases and clustered by sequence similarity. This strategy enables a more comprehensive and concise representation of small RNA populations without any data loss or data distortion. AVAILABILITY AND IMPLEMENTATION: Code and documentation of our R package at http://ibis.helmholtz-muenchen.de/deus/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

A genome-wide scan study identifies a single nucleotide substitution in MC1R gene associated with white coat colour in fallow deer (Dama dama).

BACKGROUND: The coat colour of fallow deer is highly variable and even white animals can regularly be observed in game farming and in the wild. Affected animals do not show complete albinism but rather some residual pigmentation resembling a very pale beige dilution of coat colour. The eyes and claws of the animals are pigmented. To facilitate the conservation and management of such animals, it would be helpful to know the responsible gene and causative variant. We collected 102 samples from 22 white animals and from 80 animals with wildtype coat colour. The samples came from 12 different wild flocks or game conservations located in different regions of Germany, at the border to Luxembourg and in Poland. The genomes of one white hind and her brown calf were sequenced. RESULTS: Based on a list of colour genes of the International Federation of Pigment Cell Societies ( http://www.ifpcs.org/albinism/ ), a variant in the MC1R gene (NM_174108.2:c.143 T > C) resulting in an amino acid exchange from leucine to proline at position 48 of the MC1R receptor protein (NP_776533.1:p.L48P) was identified as a likely cause of coat colour dilution. A gene test revealed that all animals of the white phenotype were of genotype CC whereas all pigmented animals were of genotype TT or TC. The study showed that 14% of the pigmented (brown or dark pigmented) animals carried the white allele. CONCLUSIONS: A genome-wide scan study led to a molecular test to determine the coat colour of fallow deer. Identification of the MC1R gene provides a deeper insight into the mechanism of dilution. The gene marker is now available for the conservation of white fallow deer in wild and farmed animals.

A genome-wide scan study identifies a single nucleotide substitution in the tyrosinase gene associated with white coat colour in a red deer (Cervus elaphus) population.

BACKGROUND: Red deer with very pale coat colour are observed sporadically. In the red deer (Cervus elaphus) population of Reinhardswald in Germany, about 5% of animals have a white coat colour that is not associated with albinism. In order to facilitate the conservation of the animals, it should be determined whether and to what extent brown animals carry the white gene. For this purpose, samples of one white hind and her brown calf were available for whole genome sequencing to identify the single nucleotide polymorphism(s) responsible for the white phenotype. Subsequently, samples from 194 brown and 11 white animals were genotyped. RESULTS: Based on a list of colour genes of the International Federation of Pigment Cell Societies, a non-synonymous mutation with exchange of a glycine residue at position 291 of the tyrosinase protein by arginine was identified as the cause of dilution of the coat colour. A gene test led to exactly matching genotypes in all examined animals. The study showed that 14% of the brown animals carry the white gene. This provides a simple and reliable way of conservation for the white animals. However, results could not be transferred to another, unrelated red deer population with white animals. Although no brown animals with a white tyrosinase genotype were detected, the cause for the white colouring in this population was different. CONCLUSIONS: A gene test for the conservation of white red deer is available for the population of the Reinhardswald. While mutations in the tyrosinase are commonly associated with oculocutaneous albinism type 1, the amino acid exchange at position 291 was found to be associated with coat colour dilution in Cervus elaphus.

A splice donor variant in CCDC189 is associated with asthenospermia in Nordic Red dairy cattle.

BACKGROUND: Cattle populations are highly amenable to the genetic mapping of male reproductive traits because longitudinal data on ejaculate quality and dense microarray-derived genotypes are available for thousands of artificial insemination bulls. Two young Nordic Red bulls delivered sperm with low progressive motility (i.e., asthenospermia) during a semen collection period of more than four months. The bulls were related through a common ancestor on both their paternal and maternal ancestry. Thus, a recessive mode of inheritance of asthenospermia was suspected. RESULTS: Both bulls were genotyped at 54,001 SNPs using the Illumina BovineSNP50 Bead chip. A scan for autozygosity revealed that they were identical by descent for a 2.98 Mb segment located on bovine chromosome 25. This haplotype was not found in the homozygous state in 8557 fertile bulls although five homozygous haplotype carriers were expected (P = 0.018). Whole genome-sequencing uncovered that both asthenospermic bulls were homozygous for a mutation that disrupts a canonical 5' splice donor site of CCDC189 encoding the coiled-coil domain containing protein 189. Transcription analysis showed that the derived allele activates a cryptic splice site resulting in a frameshift and premature termination of translation. The mutated CCDC189 protein is truncated by more than 40%, thus lacking the flagellar C1a complex subunit C1a-32 that is supposed to modulate the physiological movement of the sperm flagella. The mutant allele occurs at a frequency of 2.5% in Nordic Red cattle. CONCLUSIONS: Our study in cattle uncovered that CCDC189 is required for physiological movement of sperm flagella thus enabling active progression of spermatozoa and fertilization. A direct gene test may be implemented to monitor the asthenospermia-associated allele and prevent the birth of homozygous bulls that are infertile. Our results have been integrated in the Online Mendelian Inheritance in Animals (OMIA) database ( https://omia.org/OMIA002167/9913/ ).

Effect of plant chemical variation and mutualistic ants on the local population genetic structure of an aphid herbivore.

Plants exhibit impressive genetic and chemical diversity, not just between species but also within species, and the importance of plant intraspecific variation for structuring ecological communities is well known. When there is variation at the local population level, this can create a spatially heterogeneous habitat for specialised herbivores potentially leading to non-random distribution of individuals across host plants. Plant variation can affect herbivores directly and indirectly via a third species, resulting in variable herbivore growth rates across different host plants. Herbivores also exhibit within-species variation, with some genotypes better adapted to some plant variants than others. We genotyped aphids collected across 2 years from a field site containing ~200 patchily distributed host plants that exhibit high chemical diversity. The distribution of aphid genotypes, their ant mutualists, and other predators was assessed across the plants. We present evidence that the local distribution of aphid (Metopeurum fuscoviride) genotypes across host-plant individuals is associated with variation in the plant volatiles (chemotypes) and non-volatile metabolites (metabotypes) of their host plant tansy (Tanacetum vulgare). Furthermore, these interactions in the field were influenced by plant-host preferences of aphid-mutualist ants. Our results emphasise that plant intraspecific variation can structure ecological communities not only at the species level but also at the genetic level within species and that this effect can be enhanced through indirect interactions with a third species.

Accurate sequence variant genotyping in cattle using variation-aware genome graphs.

BACKGROUND: Genotyping of sequence variants typically involves, as a first step, the alignment of sequencing reads to a linear reference genome. Because a linear reference genome represents only a small fraction of all the DNA sequence variation within a species, reference allele bias may occur at highly polymorphic or divergent regions of the genome. Graph-based methods facilitate the comparison of sequencing reads to a variation-aware genome graph, which incorporates a collection of non-redundant DNA sequences that segregate within a species. We compared the accuracy and sensitivity of graph-based sequence variant genotyping using the Graphtyper software to two widely-used methods, i.e., GATK and SAMtools, which rely on linear reference genomes using whole-genome sequencing data from 49 Original Braunvieh cattle. RESULTS: We discovered 21,140,196, 20,262,913, and 20,668,459 polymorphic sites using GATK, Graphtyper, and SAMtools, respectively. Comparisons between sequence variant genotypes and microarray-derived genotypes showed that Graphtyper outperformed both GATK and SAMtools in terms of genotype concordance, non-reference sensitivity, and non-reference discrepancy. The sequence variant genotypes that were obtained using Graphtyper had the smallest number of Mendelian inconsistencies between sequence-derived single nucleotide polymorphisms and indels in nine sire-son pairs. Genotype phasing and imputation using the Beagle software improved the quality of the sequence variant genotypes for all the tools evaluated, particularly for animals that were sequenced at low coverage. Following imputation, the concordance between sequence- and microarray-derived genotypes was almost identical for the three methods evaluated, i.e., 99.32, 99.46, and 99.24% for GATK, Graphtyper, and SAMtools, respectively. Variant filtration based on commonly used criteria improved genotype concordance slightly but it also decreased sensitivity. Graphtyper required considerably more computing resources than SAMtools but less than GATK. CONCLUSIONS: Sequence variant genotyping using Graphtyper is accurate, sensitive and computationally feasible in cattle. Graph-based methods enable sequence variant genotyping from variation-aware reference genomes that may incorporate cohort-specific sequence variants, which is not possible with the current implementation of state-of-the-art methods that rely on linear reference genomes.

Maternal placenta modulates a deleterious fetal mutation.

Intrauterine growth restriction (IUGR) is caused by dysregulation of placental metabolism. Paternally inherited IUGR mutations in the fetus influence maternal physiology via the placenta. However, it is not known whether the maternal placenta also affects the extent of IUGR in such fetuses. In cattle and other ruminants, maternal-fetal communication occurs primarily at the placentomes. We previously identified a 3΄ deletion in the noncoding MER1 repeat containing imprinted transcript 1 (MIMT1) gene that, when inherited from the sire, causes IUGR and late abortion in Ayshire cattle with variable levels of severity. Here, we compared the transcriptome and genomic imprinting in fetal and maternal placentome components of wild-type and MIMT1Del/WT fetuses before IUGR became apparent, to identify key early events. Transcriptome analysis revealed fewer differentially expressed genes in maternal than fetal MIMT1Del/WT placentome. AST1, within the PEG3 domain, was the only gene consistently reduced in IUGR in both fetal and maternal samples. Several genes showed an imprinting pattern associated with IUGR, of which only secernin 3 (SCRN3) and paternally expressed 3 (PEG3) were differentially imprinted in both placentome components. Loss of strictly monoallelic, allele-specific expression (∼80:20) of PEG3 in the maternal MIMT1Del/WT placenta could be associated with incomplete penetrance of MIMT1Del. Our data show that dysregulation of the PEG3 domain is involved in IUGR, but also reveal that maternal placental tissues may affect the penetrance of the paternally inherited IUGR mutation.

A nonsense mutation in TMEM95 encoding a nondescript transmembrane protein causes idiopathic male subfertility in cattle.

Genetic variants underlying reduced male reproductive performance have been identified in humans and model organisms, most of them compromising semen quality. Occasionally, male fertility is severely compromised although semen analysis remains without any apparent pathological findings (i.e., idiopathic subfertility). Artificial insemination (AI) in most cattle populations requires close examination of all ejaculates before insemination. Although anomalous ejaculates are rejected, insemination success varies considerably among AI bulls. In an attempt to identify genetic causes of such variation, we undertook a genome-wide association study (GWAS). Imputed genotypes of 652,856 SNPs were available for 7962 AI bulls of the Fleckvieh (FV) population. Male reproductive ability (MRA) was assessed based on 15.3 million artificial inseminations. The GWAS uncovered a strong association signal on bovine chromosome 19 (P = 4.08 × 10(-59)). Subsequent autozygosity mapping revealed a common 1386 kb segment of extended homozygosity in 40 bulls with exceptionally poor reproductive performance. Only 1.7% of 35,671 inseminations with semen samples of those bulls were successful. None of the bulls with normal reproductive performance was homozygous, indicating recessive inheritance. Exploiting whole-genome re-sequencing data of 43 animals revealed a candidate causal nonsense mutation (rs378652941, c.483C>A, p.Cys161X) in the transmembrane protein 95 encoding gene TMEM95 which was subsequently validated in 1990 AI bulls. Immunohistochemical investigations evidenced that TMEM95 is located at the surface of spermatozoa of fertile animals whereas it is absent in spermatozoa of subfertile animals. These findings imply that integrity of TMEM95 is required for an undisturbed fertilisation. Our results demonstrate that deficiency of TMEM95 severely compromises male reproductive performance in cattle and reveal for the first time a phenotypic effect associated with genomic variation in TMEM95.

A missense mutation in TUBD1 is associated with high juvenile mortality in Braunvieh and Fleckvieh cattle.

BACKGROUND: Haplotypes with reduced or missing homozygosity may harbor deleterious alleles that compromise juvenile survival. A scan for homozygous haplotype deficiency revealed a short segment on bovine chromosome 19 (Braunvieh haplotype 2, BH2) that was associated with high juvenile mortality in Braunvieh cattle. However, the molecular genetic underpinnings and the pathophysiology of BH2 remain to be elucidated. RESULTS: The frequency of BH2 was 6.5 % in 8,446 Braunvieh animals from the national bovine genome databases. Both perinatal and juvenile mortality of BH2 homozygous calves were higher than the average in Braunvieh cattle resulting in a depletion of BH2 homozygous adult animals (P = 9.3x10(-12)). The analysis of whole-genome sequence data from 54 Braunvieh animals uncovered a missense mutation in TUBD1 (rs383232842, p.H210R) that was compatible with recessive inheritance of BH2. The availability of sequence data of 236 animals from diverse bovine populations revealed that the missense mutation also segregated at a low frequency (1.7 %) in the Fleckvieh breed. A validation study in 37,314 Fleckvieh animals confirmed high juvenile mortality of homozygous calves (P = 2.2x10(-15)). Our findings show that the putative disease allele is located on an ancestral haplotype that segregates in Braunvieh and Fleckvieh cattle. To unravel the pathophysiology of BH2, six homozygous animals were examined at the animal clinic. Clinical and pathological findings revealed that homozygous calves suffered from chronic airway disease possibly resulting from defective cilia in the respiratory tract. CONCLUSIONS: A missense mutation in TUBD1 is associated with high perinatal and juvenile mortality in Braunvieh and Fleckvieh cattle. The mutation is located on a common haplotype likely originating from an ancient ancestor of Braunvieh and Fleckvieh cattle. Our findings demonstrate for the first time that deleterious alleles may segregate across closed cattle breeds without recent admixture. Homozygous calves suffer from chronic airway disease resulting in poor growth performance and high juvenile mortality. The respiratory manifestations resemble key features of diseases resulting from impaired function of airway cilia.

A nonsense mutation in the COL7A1 gene causes epidermolysis bullosa in Vorderwald cattle.

BACKGROUND: The widespread use of individual sires for artificial insemination promotes the propagation of recessive conditions. Inadvertent matings between unnoticed carriers of deleterious alleles may result in the manifestation of fatal phenotypes in their progeny. Breeding consultants and farmers reported on Vorderwald calves with a congenital skin disease. The clinical findings in affected calves were compatible with epidermolysis bullosa. RESULTS: Pedigree analysis indicated autosomal recessive inheritance of epidermolysis bullosa in Vorderwald cattle. We genotyped two diseased and 41 healthy animals at 41,436 single nucleotide polymorphisms and performed whole-genome haplotype-based association testing, which allowed us to map the locus responsible for the skin disease to the distal end of bovine chromosome 22 (P = 8.0 × 10-14). The analysis of whole-genome re-sequencing data of one diseased calf, three obligate mutation carriers and 1682 healthy animals from various bovine breeds revealed a nonsense mutation (rs876174537, p.Arg1588X) in the COL7A1 gene that segregates with the disease. The same mutation was previously detected in three calves with dystrophic epidermolysis bullosa from the Rotes Höhenvieh cattle breed. We show that diseased animals from Vorderwald and Rotes Höhenvieh cattle are identical by descent for an 8.72 Mb haplotype encompassing rs876174537 indicating they inherited the deleterious allele from a recent common ancestor. CONCLUSIONS: Autosomal recessive epidermolysis bullosa in Vorderwald and Rotes Höhenvieh cattle is caused by a nonsense mutation in the COL7A1 gene. Our findings demonstrate that deleterious alleles may segregate across cattle populations without apparent admixture. The identification of the causal mutation now enables the reliable detection of carrier animals. Genome-based mating strategies can avoid inadvertent matings of carrier animals thereby preventing the birth of homozygous calves that suffer from a painful skin disease.

A frameshift mutation in ARMC3 is associated with a tail stump sperm defect in Swedish Red (Bos taurus) cattle.

BACKGROUND: Artificial insemination is widely used in many cattle breeding programs. Semen samples of breeding bulls are collected and closely examined immediately after collection at artificial insemination centers. Only ejaculates without anomalous findings are retained for artificial insemination. Although morphological aberrations of the spermatozoa are a frequent reason for discarding ejaculates, the genetic determinants underlying poor semen quality are scarcely understood. RESULTS: A tail stump sperm defect was observed in three bulls of the Swedish Red cattle breed. The spermatozoa of affected bulls were immotile because of severely disorganized tails indicating disturbed spermatogenesis. We genotyped three affected bulls and 18 unaffected male half-sibs at 46,035 SNPs and performed homozygosity mapping to map the fertility disorder to an 8.42 Mb interval on bovine chromosome 13. The analysis of whole-genome re-sequencing data of an affected bull and 300 unaffected animals from eleven cattle breeds other than Swedish Red revealed a 1 bp deletion (Chr13: 24,301,425 bp, ss1815612719) in the eleventh exon of the armadillo repeat containing 3-encoding gene (ARMC3) that was compatible with the supposed recessive mode of inheritance. The deletion is expected to alter the reading frame and to induce premature translation termination (p.A451fs26). The mutated protein is shortened by 401 amino acids (46 %) and lacks domains that are likely essential for normal protein function. CONCLUSIONS: We report the phenotypic and genetic characterization of a sterilizing tail stump sperm defect in the Swedish Red cattle breed. Exploiting high-density genotypes and massive re-sequencing data enabled us to identify the most likely causal mutation for the fertility disorder in bovine ARMC3. Our results provide the basis for monitoring the mutated variant in the Swedish Red cattle population and for the early identification of infertile animals.

A frameshift mutation in GON4L is associated with proportionate dwarfism in Fleckvieh cattle.

BACKGROUND: Low birth weight and postnatal growth restriction are the most evident symptoms of dwarfism. Accompanying skeletal aberrations may compromise the general condition and locomotion of affected individuals. Several paternal half-sibs with a low birth weight and a small size were born in 2013 in the Fleckvieh cattle population. RESULTS: Affected calves were strikingly underweight at birth in spite of a normal gestation length and had craniofacial abnormalities such as elongated narrow heads and brachygnathia inferior. In spite of a normal general condition, their growth remained restricted during rearing. We genotyped 27 affected and 10,454 unaffected animals at 44,672 single nucleotide polymorphisms and performed association tests followed by homozygosity mapping, which allowed us to map the locus responsible for growth failure to a 1.85-Mb segment on bovine chromosome 3. Analysis of whole-genome re-sequencing data from one affected and 289 unaffected animals revealed a 1-bp deletion (g.15079217delC, rs723240647) in the coding region of the GON4L gene that segregated with the dwarfism-associated haplotype. We showed that the deletion induces intron retention and premature termination of translation, which can lead to a severely truncated protein that lacks domains that are likely essential to normal protein function. The widespread use of an undetected carrier bull for artificial insemination has resulted in a tenfold increase in the frequency of the deleterious allele in the female population. CONCLUSIONS: A frameshift mutation in GON4L is associated with autosomal recessive proportionate dwarfism in Fleckvieh cattle. The mutation has segregated in the population for more than 50 years without being recognized as a genetic disorder. However, the widespread use of an undetected carrier bull for artificial insemination caused a sudden accumulation of homozygous calves with dwarfism. Our findings provide the basis for genome-based mating strategies to avoid the inadvertent mating of carrier animals and thereby prevent the birth of homozygous calves with impaired growth.

Homozygous haplotype deficiency reveals deleterious mutations compromising reproductive and rearing success in cattle.

BACKGROUND: Cattle breeding populations are susceptible to the propagation of recessive diseases. Individual sires generate tens of thousands of progeny via artificial insemination. The frequency of deleterious alleles carried by such sires may increase considerably within few generations. Deleterious alleles manifest themselves often by missing homozygosity resulting from embryonic/fetal, perinatal or juvenile lethality of homozygotes. RESULTS: A scan for homozygous haplotype deficiency in 25,544 Fleckvieh cattle uncovered four haplotypes affecting reproductive and rearing success. Exploiting whole-genome resequencing data from 263 animals facilitated to pinpoint putatively causal mutations in two of these haplotypes. A mutation causing an evolutionarily unlikely substitution in SUGT1 was perfectly associated with a haplotype compromising insemination success. The mutation was not found in homozygous state in 10,363 animals (P=1.79×10(-5)) and is thus likely to cause lethality of homozygous embryos. A frameshift mutation in SLC2A2 encoding glucose transporter 2 (GLUT2) compromises calf survival. The mutation leads to premature termination of translation and activates cryptic splice sites resulting in multiple exon variants also with premature translation termination. The affected calves exhibit stunted growth, resembling the phenotypic appearance of Fanconi-Bickel syndrome in humans (OMIM 227810), which is also caused by mutations in SLC2A2. CONCLUSIONS: Exploiting comprehensive genotype and sequence data enabled us to reveal two deleterious alleles in SLC2A2 and SUGT1 that compromise pre- and postnatal survival in homozygous state. Our results provide the basis for genome-assisted approaches to avoiding inadvertent carrier matings and to improving reproductive and rearing success in Fleckvieh cattle.