207 research outputs found
Lineage-specific evolution of the vertebrate Otopetrin gene family revealed by comparative genomic analyses
No full textGrupo de trabajo: NISC Comparative Sequencing Program. -- 18 páginas, 6 figuras, 2 tablas.[Background]: Mutations in the Otopetrin 1 gene (Otop1) in mice and fish produce an unusual bilateral vestibular pathology that involves the absence of otoconia without hearing impairment. The encoded protein, Otop1, is the only functionally characterized member of the Otopetrin Domain Protein (ODP) family; the extended sequence and structural preservation of ODP proteins in metazoans suggest a conserved functional role. Here, we use the tools of sequence- and cytogenetic-based comparative genomics to study the Otop1 and the Otop2-Otop3 genes and to establish their genomic context in 25 vertebrates. We extend our evolutionary study to include the gene mutated in Usher syndrome (USH) subtype 1G (Ush1g), both because of the head-to-tail clustering of Ush1g with Otop2 and because Otop1 and Ush1g mutations result in inner ear phenotypes.[Results]: We established that OTOP1 is the boundary gene of an inversion polymorphism on human chromosome 4p16 that originated in the common human-chimpanzee lineage more than 6 million years ago. Other lineagespecific evolutionary events included a three-fold expansion of the Otop genes in Xenopus tropicalis and of Ush1g in teleostei fish. The tight physical linkage between Otop2 and Ush1g is conserved in all vertebrates. To further understand the functional organization of the Ushg1-Otop2 locus, we deduced a putative map of binding sites for CCCTC-binding factor (CTCF), a mammalian insulator transcription factor, from genome-wide chromatin immunoprecipitation-sequencing (ChIP-seq) data in mouse and human embryonic stem (ES) cells combined with detection of CTCF-binding motifs.[Conclusions]: The results presented here clarify the evolutionary history of the vertebrate Otop and Ush1g families, and establish a framework for studying the possible interaction(s) of Ush1g and Otop in developmental pathways.This work was supported by the NHGRI Intramural Research Program, NIDCD grant DC02236 (DMO); and NHGRI grants HG002385 and HG0058815 (EEE). EEE is an investigator of the Howard Hughes Medical Institute.Peer Reviewe
Megalencephaly syndromes: exome pipeline strategies for detecting low-level mosaic mutations
No full textTwo megalencephaly (MEG) syndromes, megalencephaly-capillary malformation (MCAP) and megalencephaly-polymicrogyriapolydactyly?-hydrocephalus(MPPH), have recently been defined on the basis of physical and neuroimaging features. Subsequently, exome sequencing of ten MEG cases identified de-novo postzygotic mutations in PIK3CA which cause MCAP and de-novo mutations in AKT and PIK3R2 which cause MPPH. Here we present findings from exome sequencing three unrelated megalencephaly patients which identified a causal PIK3CA mutation in two cases and a causal PIK3R2 mutation in the third case. However, our patient with the PIK3R2 mutation which is considered to cause MPPH has a marked bifrontal band heterotopia which is a feature of MCAP. Furthermore, one of our patients with a PIK3CA mutation lacks syndactyly/polydactyly which is a characteristic of MCAP. These findings suggest that the overlap between MCAP and MPPH may be greater than the available studies suggest. In addition, the PIK3CA mutation in one of our patients could not be detected using standard exome analysis because the mutation was observed at a low frequency consistent with somatic mosaicism. We have therefore investigated several alternative methods of exome analysis and demonstrate that alteration of the initial allele frequency spectrum (AFS), used as a prior for variant calling in samtools, had the greatest power to detect variants with low mutant allele frequencies in our 3 MEG exomes and in simulated data. We therefore recommend non-default settings of the AFS in combination with stringent quality control when searching for causal mutation(s) that could have low levels of mutant reads due to post-zygotic mutatio
Comparative BAC-based mapping in the white-throated sparrow, a novel behavioral genomics model, using interspecies overgo hybridization
No full textBACKGROUND
The genomics era has produced an arsenal of resources from sequenced organisms allowing researchers to target species that do not have comparable mapping and sequence information. These new "non-model" organisms offer unique opportunities to examine environmental effects on genomic patterns and processes. Here we use comparative mapping as a first step in characterizing the genome organization of a novel animal model, the white-throated sparrow (Zonotrichia albicollis), which occurs as white or tan morphs that exhibit alternative behaviors and physiology. Morph is determined by the presence or absence of a complex chromosomal rearrangement. This species is an ideal model for behavioral genomics because the association between genotype and phenotype is absolute, making it possible to identify the genomic bases of phenotypic variation.
FINDINGS
We initiated a genomic study in this species by characterizing the white-throated sparrow BAC library via filter hybridization with overgo probes designed for the chicken, turkey, and zebra finch. Cross-species hybridization resulted in 640 positive sparrow BACs assigned to 77 chicken loci across almost all macro-and microchromosomes, with a focus on the chromosomes associated with morph. Out of 216 overgos, 36% of the probes hybridized successfully, with an average number of 3.0 positive sparrow BACs per overgo.
CONCLUSIONS
These data will be utilized for determining chromosomal architecture and for fine-scale mapping of candidate genes associated with phenotypic differences. Our research confirms the utility of interspecies hybridization for developing comparative maps in other non-model organisms
The Hox-TALE has been wagging for a long time
No full textHox and TALE proteins interact in a sea anemone, just as they do in flies and mice, indicating that the Hox-TALE system originated very early in animal evolution.Peer reviewe
Genome sequencing defines phylogeny and spread of methicillin-resistant Staphylococcus aureus in a high transmission setting.
No full textMethicillin-resistant Staphylococcus aureus (MRSA) is a major cause of nosocomial infection. Whole-genome sequencing of MRSA has been used to define phylogeny and transmission in well-resourced healthcare settings, yet the greatest burden of nosocomial infection occurs in resource-restricted settings where barriers to transmission are lower. Here, we study the flux and genetic diversity of MRSA on ward and individual patient levels in a hospital where transmission was common. We repeatedly screened all patients on two intensive care units for MRSA carriage over a 3-mo period. All MRSA belonged to multilocus sequence type 239 (ST 239). We defined the population structure and charted the spread of MRSA by sequencing 79 isolates from 46 patients and five members of staff, including the first MRSA-positive screen isolates and up to two repeat isolates where available. Phylogenetic analysis identified a flux of distinct ST 239 clades over time in each intensive care unit. In total, five main clades were identified, which varied in the carriage of plasmids encoding antiseptic and antimicrobial resistance determinants. Sequence data confirmed intra- and interwards transmission events and identified individual patients who were colonized by more than one clade. One patient on each unit was the source of numerous transmission events, and deep sampling of one of these cases demonstrated colonization with a "cloud" of related MRSA variants. The application of whole-genome sequencing and analysis provides novel insights into the transmission of MRSA in under-resourced healthcare settings and has relevance to wider global health
Quantifying single nucleotide variant detection sensitivity in exome sequencing
No full textBACKGROUND: The targeted capture and sequencing of genomic regions has rapidlydemonstrated its utility in genetic studies. Inherent in this technology isconsiderable heterogeneity of target coverage and this is expected tosystematically impact our sensitivity to detect genuine polymorphisms. To fullyinterpret the polymorphisms identified in a genetic study it is often essentialto both detect polymorphisms and to understand where and with what probabilityreal polymorphisms may have been missed.RESULTS: Using down-sampling of 30 deeply sequenced exomes and a set ofgold-standard single nucleotide variant (SNV) genotype calls for each sample, we developed an empirical model relating the read depth at a polymorphic site to theprobability of calling the correct genotype at that site. We find that measuredsensitivity in SNV detection is substantially worse than that predicted from the naive expectation of sampling from a binomial. This calibrated model allows us toproduce single nucleotide resolution SNV sensitivity estimates which can bemerged to give summary sensitivity measures for any arbitrary partition of thetarget sequences (nucleotide, exon, gene, pathway, exome). These metrics aredirectly comparable between platforms and can be combined between samples to give"power estimates" for an entire study. We estimate a local read depth of 13X isrequired to detect the alleles and genotype of a heterozygous SNV 95% of thetime, but only 3X for a homozygous SNV. At a mean on-target read depth of 20X,commonly used for rare disease exome sequencing studies, we predict 5-15% ofheterozygous and 1-4% of homozygous SNVs in the targeted regions will be missed.CONCLUSIONS: Non-reference alleles in the heterozygote state have a high chanceof being missed when commonly applied read coverage thresholds are used despitethe widely held assumption that there is good polymorphism detection at thesecoverage levels. Such alleles are likely to be of functional importance inpopulation based studies of rare diseases, somatic mutations in cancer andexplaining the "missing heritability" of quantitative traits.<br/
Sequence diversity of Pan troglodytes subspecies and the impact of WFDC6 selective constraints in reproductive immunity
No full textBalancing Selection Maintains a Form of ERAP2 that Undergoes Nonsense-Mediated Decay and Affects Antigen Presentation
No full textA remarkable characteristic of the human major histocompatibility complex (MHC) is its extreme genetic diversity, which is maintained by balancing selection. In fact, the MHC complex remains one of the best-known examples of natural selection in humans, with well-established genetic signatures and biological mechanisms for the action of selection. Here, we present genetic and functional evidence that another gene with a fundamental role in MHC class I presentation, endoplasmic reticulum aminopeptidase 2 (ERAP2), has also evolved under balancing selection and contains a variant that affects antigen presentation. Specifically, genetic analyses of six human populations revealed strong and consistent signatures of balancing selection affecting ERAP2. This selection maintains two highly differentiated haplotypes (Haplotype A and Haplotype B), with frequencies 0.44 and 0.56, respectively. We found that ERAP2 expressed from Haplotype B undergoes differential splicing and encodes a truncated protein, leading to nonsense-mediated decay of the mRNA. To investigate the consequences of ERAP2 deficiency on MHC presentation, we correlated surface MHC class I expression with ERAP2 genotypes in primary lymphocytes. Haplotype B homozygotes had lower levels of MHC class I expressed on the surface of B cells, suggesting that naturally occurring ERAP2 deficiency affects MHC presentation and immune response. Interestingly, an ERAP2 paralog, endoplasmic reticulum aminopeptidase 1 (ERAP1), also shows genetic signatures of balancing selection. Together, our findings link the genetic signatures of selection with an effect on splicing and a cellular phenotype. Although the precise selective pressure that maintains polymorphism is unknown, the demonstrated differences between the ERAP2 splice forms provide important insights into the potential mechanism for the action of selection
Evolutionary dynamism of the primate LRRC37 gene family
No full textCore duplicons in the human genome represent ancestral duplication modules shared by the majority of intrachromosomal duplication blocks within a given chromosome. These cores are associated with the emergence of novel gene families in the hominoid lineage, but their genomic organization and gene characterization among other primates are largely unknown. Here, we investigate the genomic organization and expression of the core duplicon on chromosome 17 that led to the expansion of LRRC37 during primate evolution. A comparison of the LRRC37 gene family organization in human, orangutan, macaque, marmoset, and lemur genomes shows the presence of both orthologous and species-specific gene copies in all primate lineages. Expression profiling in mouse, macaque, and human tissues reveals that the ancestral expression of LRRC37 was restricted to the testis. In the hominid lineage, the pattern of LRRC37 became increasingly ubiquitous, with significantly higher levels of expression in the cerebellum and thymus, and showed a remarkable diversity of alternative splice forms. Transfection studies in HeLa cells indicate that the human FLAG-tagged recombinant LRRC37 protein is secreted after cleavage of a transmembrane precursor and its overexpression can induce filipodia formation. needs and human health
A Novel null homozygous mutation confirms <i>CACNA2D2</i> as a gene mutated in epileptic encephalopathy
No full textContribution to epileptic encephalopathy (EE) of mutations in CACNA2D2, encoding α2δ-2 subunit of Voltage Dependent Calcium Channels, is unclear. To date only one CACNA2D2 mutation altering channel functionality has been identified in a single family. In the same family, a rare CELSR3 polymorphism also segregated with disease. Involvement of CACNA2D2 in EE
is therefore not confirmed, while that of CELSR3 is questionable. In a patient with epilepsy, dyskinesia, cerebellar atrophy, psychomotor delay and dysmorphic features, offspring to consanguineous parents, we performed whole exome sequencing (WES) for homozygosity mapping and mutation detection. WES identified extended autozygosity on
chromosome 3, containing two novel homozygous candidate mutations: c.1295delA (p.Asn432fs) in CACNA2D2 and
c.G6407A (p.Gly2136Asp) in CELSR3. Gene prioritization pointed to CACNA2D2 as the most prominent candidate gene. The WES finding in CACNA2D2 resulted to be statistically significant (p = 0.032), unlike that in CELSR3. CACNA2D2 homozygous c.1295delA essentially abolished α2δ-2 expression. In summary, we identified a novel null CACNA2D2 mutation associated to a clinical phenotype strikingly similar to the Cacna2d2 null mouse model. Molecular and statistical analyses together argued
in favor of a causal contribution of CACNA2D2 mutations to EE, while suggested that finding in CELSR3, although potentially damaging, is likely incidental
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