ABSTRACT
Spermatozoa harbour a complex and environment-sensitive pool of small non-coding RNAs (sncRNAs)1, which influences offspring development and adult phenotypes1-7. Whether spermatozoa in the epididymis are directly susceptible to environmental cues is not fully understood8. Here we used two distinct paradigms of preconception acute high-fat diet to dissect epididymal versus testicular contributions to the sperm sncRNA pool and offspring health. We show that epididymal spermatozoa, but not developing germ cells, are sensitive to the environment and identify mitochondrial tRNAs (mt-tRNAs) and their fragments (mt-tsRNAs) as sperm-borne factors. In humans, mt-tsRNAs in spermatozoa correlate with body mass index, and paternal overweight at conception doubles offspring obesity risk and compromises metabolic health. Sperm sncRNA sequencing of mice mutant for genes involved in mitochondrial function, and metabolic phenotyping of their wild-type offspring, suggest that the upregulation of mt-tsRNAs is downstream of mitochondrial dysfunction. Single-embryo transcriptomics of genetically hybrid two-cell embryos demonstrated sperm-to-oocyte transfer of mt-tRNAs at fertilization and suggested their involvement in the control of early-embryo transcription. Our study supports the importance of paternal health at conception for offspring metabolism, shows that mt-tRNAs are diet-induced and sperm-borne and demonstrates, in a physiological setting, father-to-offspring transfer of sperm mitochondrial RNAs at fertilization.
Subject(s)
Diet, High-Fat , Epigenesis, Genetic , Mitochondria , RNA, Mitochondrial , Spermatozoa , Animals , Female , Humans , Male , Mice , Body Mass Index , Diet, High-Fat/adverse effects , Embryo, Mammalian/cytology , Embryo, Mammalian/embryology , Embryo, Mammalian/metabolism , Epididymis/cytology , Epigenesis, Genetic/genetics , Fertilization/genetics , Gene Expression Profiling , Gene Expression Regulation, Developmental , Mice, Inbred C57BL , Mitochondria/genetics , Mitochondria/metabolism , Mitochondria/pathology , Obesity/genetics , Obesity/metabolism , Obesity/etiology , Oocytes/metabolism , Overweight/genetics , Overweight/metabolism , Paternal Inheritance/genetics , RNA, Mitochondrial/genetics , RNA, Mitochondrial/metabolism , RNA, Small Untranslated/genetics , RNA, Small Untranslated/metabolism , RNA, Transfer/genetics , RNA, Transfer/metabolism , Spermatozoa/metabolism , Testis/cytology , Transcription, GeneticABSTRACT
STUDY QUESTION: Does mitochondrial DNA (mtDNA) diversity in modern human populations potentially pose a challenge, via mtDNA segregation, to mitochondrial replacement therapies? SUMMARY ANSWER: The magnitude of mtDNA diversity in modern human populations is as high as in mammalian model systems where strong mtDNA segregation is observed; consideration of haplotype pairs and/or haplotype matching can help avoid these potentially deleterious effects. WHAT IS KNOWN ALREADY: In mammalian models, substantial proliferative differences are observed between different mtDNA haplotypes in cellular admixtures, with larger proliferative differences arising from more diverse haplotype pairings. If maternal mtDNA is 'carried over' in human gene therapies, these proliferative differences could lead to its amplification in the resulting offspring, potentially leading to manifestation of the disease that the therapy was designed to avoid-but existing studies have not investigated whether mtDNA diversity in modern human populations is sufficient to permit significant amplification. STUDY DESIGN, SIZE, DURATION: This theoretical study used over 7500 human mtDNA sequences from The National Center for Biotechnology Information (NCBI), a range of international and British mtDNA surveys, and 2011 census data. PARTICIPANTS/MATERIALS, SETTING, METHODS: A stochastic simulation approach was used to model random haplotype pairings from within different regions. In total, 1000 simulated pairings were analysed using the basic local alignment search tool (BLAST) for each region. Previous data from mouse models were used to estimate proliferative differences. MAIN RESULTS AND THE ROLE OF CHANCE: Even within the same haplogroup, differences of around 20-80 single-nucleotide polymorphisms (SNPs) are common between mtDNAs admixed in random pairings. These values are sufficient to lead to substantial segregation in mouse models over an organismal lifetime, even given low starting heteroplasmy, inducing increases from 5% to 35% over 1 year. Substantial population mixing in modern UK cities increases the expected genetic differences. Hence, the likely genetic differences between humans randomly sampled from a population may well allow substantial amplification of a disease-carrying mtDNA haplotype over the timescale of a human lifetime. We report ranges and mean differences for all statistics to quantify uncertainty in our results. LIMITATIONS/REASONS FOR CAUTION: The mapping from mouse and other mammalian models to the human system is challenging, as timescales and mechanisms may differ. Reporting biases in NCBI mtDNA data, if present, may affect the statistics we compute. We discuss the robustness of our findings in the light of these concerns. WIDER IMPLICATIONS OF THE FINDINGS: Matching the mtDNA haplotypes of the mother and third-party donor in mitochondrial replacement therapies is supported as a means of ameliorating the potentially deleterious results of human mtDNA diversity. We present a chart of expected SNP differences between mtDNA haplogroups, allowing the selection of optimal partners for therapies. LARGE SCALE DATA: N/A STUDY FUNDING/COMPETING INTERESTS: The authors report no external funding sources or conflicts of interest.
Subject(s)
DNA, Mitochondrial/genetics , Genetic Therapy/methods , Haplotypes/genetics , Humans , Mitochondria/genetics , Models, Biological , Polymorphism, Single Nucleotide/geneticsABSTRACT
REASONS FOR PERFORMING THE STUDY: Equine hoof canker is a chronic proliferative pododermatitis of as yet unknown aetiology. Like equine sarcoid disease, canker is a therapy-resistant disorder characterised by hyperkeratosis, acanthosis and a marked tendency to recur. HYPOTHESIS: There is an association of sarcoid-inducing bovine papillomaviruses of types 1 and 2 (BPV-1, BPV-2) with hoof canker disease. METHODS: Using PCR-based techniques, we assessed canker tissue, intact skin and/or peripheral blood mononuclear cells (PBMCs) of 25 canker-affected horses for the presence of sarcoid-associated BPV-1 and -2. RESULTS: Conventional PCR revealed BPV-1/-2 DNA in 24/24 canker, 12/13 skin and 10/11 PBMC DNA isolates. Using inverse PCR, full-length BPV episomes were detected in 1/5 canker specimens. Sequencing of viral early and late genes amplified from canker, intact skin and PBMC DNA of 2 cases revealed an overall identity of 98% to BPV-1. Viral DNA loads amounted to ≤16 copies per cell in canker tissue and intact skin, and to ≤0.35 copies per PBMC, as determined by quantitative PCR. Using RT-PCR, the viral major oncogene E5 was shown to be transcribed in 2/4 canker tissue specimens and 5/7 PBMC isolates. Immunocapture PCR from 7 canker and 6 skin extract supernatants revealed capsomere-associated viral DNA in one canker and one skin sample. Hoof tissue, skin and PBMCs collected from 13 individuals with no signs of canker or BPV-related malignancies scored negative throughout the experiments. CONCLUSION: These findings suggest that the observed presence of BPV-1/-2 in canker-affected horses is not coincidental but indicative of an active contribution to hoof canker disease. POTENTIAL RELEVANCE: The use of antivirals and/or immune modulators may help improving canker therapy.
Subject(s)
Bovine papillomavirus 1/isolation & purification , Foot Diseases/veterinary , Hoof and Claw/virology , Horse Diseases/virology , Papillomavirus Infections/veterinary , Skin/virology , Amino Acid Sequence , Animals , DNA, Viral/isolation & purification , Foot Diseases/virology , Horses , Leukocytes, Mononuclear/virology , Papillomavirus Infections/virology , Viral Proteins/chemistry , Viral Proteins/isolation & purificationABSTRACT
In February 2008 an epidemiological field study on arthropod-borne infections in dogs was carried out in Praia, the capital city of Cape Verde. For this purpose 130 dogs were included in the study. Of these, 94.6% were infested with ticks. Altogether, 1293 ticks of the genus Rhipicephalus (in all evaluated cases R. sanguineus) were collected. Examination for haemotropic parasites was performed via polymerase chain reaction (PCR). Lymph node fine-needle aspirates were screened by PCR for Leishmania infantum infections in 20 dogs with enlarged lymph nodes. Our investigation revealed two species of protozoa (Babesia canis vogeli and Hepatozoon canis) and two species of rickettsiae (Anaplasma platys and Ehrlichia canis). In 101 dogs (77.7%) DNA of one or more pathogens was detected. The PCR examination for H. canis was positive in 83 dogs (63.8%), for E. canis in 34 dogs (26.2%), for A. platys in 10 dogs (7.7%) and for B. canis in five dogs (3.8%), whereas neither B. gibsoni nor L. infantum DNA could be detected. Of the infected dogs, 71.3% had a monoinfection, 27.7% had infections with two pathogens and 1.0% with three pathogens. B. canis, H. canis, E. canis, A. platys and their vector tick R. sanguineus are endemic to Cape Verde and can be present in dogs in high prevalences. These results outline the risk of importing tropical canine diseases when Capeverdian stray dogs are taken to Europe.