Many existing cohorts contain a range of relatedness between genotyped individuals, either by design or by chance. Haplotype estimation in such cohorts is a central step in many downstream analyses. ...Using genotypes from six cohorts from isolated populations and two cohorts from non-isolated populations, we have investigated the performance of different phasing methods designed for nominally 'unrelated' individuals. We find that SHAPEIT2 produces much lower switch error rates in all cohorts compared to other methods, including those designed specifically for isolated populations. In particular, when large amounts of IBD sharing is present, SHAPEIT2 infers close to perfect haplotypes. Based on these results we have developed a general strategy for phasing cohorts with any level of implicit or explicit relatedness between individuals. First SHAPEIT2 is run ignoring all explicit family information. We then apply a novel HMM method (duoHMM) to combine the SHAPEIT2 haplotypes with any family information to infer the inheritance pattern of each meiosis at all sites across each chromosome. This allows the correction of switch errors, detection of recombination events and genotyping errors. We show that the method detects numbers of recombination events that align very well with expectations based on genetic maps, and that it infers far fewer spurious recombination events than Merlin. The method can also detect genotyping errors and infer recombination events in otherwise uninformative families, such as trios and duos. The detected recombination events can be used in association scans for recombination phenotypes. The method provides a simple and unified approach to haplotype estimation, that will be of interest to researchers in the fields of human, animal and plant genetics.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The CRISPR/Cas9 bacterial system has proven to be an powerful tool for genetic manipulation in several organisms, but the efficiency of sequence replacement by homologous direct repair (HDR) is ...substantially lower than random indel creation. Many studies focused on improving HDR efficiency using double sgRNA, cell synchronization cycle, and the delivery of single-stranded oligo DNA nucleotides (ssODN) with a rational design. In this study, we evaluate these three methods' synergistic effects to improve HDR efficiency. For our tests, we have chosen the TNFα gene (NM_000594) for its crucial role in various biological processes and diseases. For the first time, our results showed how the use of two sgRNA with asymmetric donor design and triple transfection events dramatically increase the HDR efficiency from an undetectable HDR event to 39% of HDR efficiency and provide a new strategy to facilitate CRISPR/Cas9-mediated human genome editing. Besides, we demonstrated that the TNFα locus could be edited with CRISPR/Cas9 methodology, an opportunity to safely correct, in the future, the specific mutations of each patient.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The release of circulating tumor cells (CTCs) into vasculature is an early event in the metastatic process. The analysis of CTCs in patients has recently received widespread attention because of its ...clinical implications, particularly for precision medicine. Accumulated evidence documents a large heterogeneity in CTCs across patients. Currently, the most accepted view is that tumor cells with an intermediate phenotype between epithelial and mesenchymal have the highest plasticity. Indeed, the existence of a meta-stable or partial epithelial⁻mesenchymal transition (EMT) cell state, with both epithelial and mesenchymal features, can be easily reconciled with the concept of a highly plastic stem-like state. A close connection between EMT and cancer stem cells (CSC) traits, with enhanced metastatic competence and drug resistance, has also been described. Accordingly, a subset of CTCs consisting of CSC, present a stemness profile, are able to survive chemotherapy, and generate metastases after xenotransplantation in immunodeficient mice. In the present review, we discuss the current evidence connecting CTCs, EMT, and stemness. An improved understanding of the CTC/EMT/CSC connections may uncover novel therapeutic targets, irrespective of the tumor type, since most cancers seem to harbor a pool of CSCs, and disclose important mechanisms underlying tumorigenicity.
Bitter molecules in humans are detected by ∼25 G protein-coupled receptors (GPCRs). The lack of atomic resolution structure for any of them is complicating an in depth understanding of the molecular ...mechanisms underlying bitter taste perception. Here, we investigate the molecular determinants of the interaction of the TAS2R38 bitter taste receptor with its agonists phenylthiocarbamide (PTC) and propylthiouracil (PROP). We use the recently developed hybrid Molecular Mechanics/Coarse Grained (MM/CG) method tailored specifically for GPCRs. The method, through an extensive exploration of the conformational space in the binding pocket, allows the identification of several residues important for agonist binding that would have been very difficult to capture from the standard bioinformatics/docking approach. Our calculations suggest that both agonists bind to Asn103, Phe197, Phe264 and Trp201, whilst they do not interact with the so-called extra cellular loop 2, involved in cis-retinal binding in the GPCR rhodopsin. These predictions are consistent with data sets based on more than 20 site-directed mutagenesis and functional calcium imaging experiments of TAS2R38. The method could be readily used for other GPCRs for which experimental information is currently lacking.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Campi Flegrei is an active, resurgent caldera that is located a few kilometres west of the city of Naples, a densely populated urban settlement in southern Italy. Identifying, locating at depth and ...better defining the geometry of the magma feeding system of the caldera is highly relevant for assessing and monitoring its volcanic hazard. Based on a high resolution seismic reflection data set, we investigated the deep structure of the volcano. Here we show that seismic wave amplitude variations with distance from the radiating source provide clear evidence for large amplitude seismic reflections from the top of an extended supercritical fluid‐bearing rock formation at about 3,000 m and of an about 7,500 m deep, 1,000 m thick, low velocity layer, which is associated with a mid‐crust, partial melting zone beneath the caldera. The modeling of magma properties based on measured seismic velocities indicates a relatively high melt percentage (in the range 80–90%). These new data suggest that a large magmatic sill is present well within the basement formations, which is possibly linked to the surface through a system of deep fractures bordering the caldera. The lateral extension and similar depth of the melt zone observed beneath the nearby Mt.Vesuvius support the hypothesis of a single continuous magma reservoir feeding both of these volcanoes.
Diverse genetic backgrounds often lead to phenotypic heterogeneity in cardiomyopathies (CMPs). Previous genotype-phenotype studies have primarily focused on the analysis of a single phenotype, and ...the diagnostic and prognostic features of the CMP genotype across different phenotypic expressions remain poorly understood.
We sought to define differences in outcome prediction when stratifying patients based on phenotype at presentation compared with genotype in a large cohort of patients with CMPs and positive genetic testing.
Dilated cardiomyopathy (DCM), arrhythmogenic right ventricular cardiomyopathy, left-dominant arrhythmogenic cardiomyopathy, and biventricular arrhythmogenic cardiomyopathy were examined in this study. A total of 281 patients (80% DCM) with pathogenic or likely pathogenic variants were included. The primary and secondary outcomes were: 1) all-cause mortality (D)/heart transplant (HT); 2) sudden cardiac death/major ventricular arrhythmias (SCD/MVA); and 3) heart failure-related death (DHF)/HT/left ventricular assist device implantation (LVAD).
Survival analysis revealed that SCD/MVA events occurred more frequently in patients without a DCM phenotype and in carriers of DSP, PKP2, LMNA, and FLNC variants. However, after adjustment for age and sex, genotype-based classification, but not phenotype-based classification, was predictive of SCD/MVA. LMNA showed the worst trends in terms of D/HT and DHF/HT/LVAD.
Genotypes were associated with significant phenotypic heterogeneity in genetic cardiomyopathies. Nevertheless, in our study, genotypic-based classification showed higher precision in predicting the outcome of patients with CMP than phenotype-based classification. These findings add to our current understanding of inherited CMPs and contribute to the risk stratification of patients with positive genetic testing.
We present the results of a GWAS of food liking conducted on 161,625 participants from the UK-Biobank. Liking was assessed over 139 specific foods using a 9-point scale. Genetic correlations coupled ...with structural equation modelling identified a multi-level hierarchical map of food-liking with three main dimensions: "Highly-palatable", "Acquired" and "Low-caloric". The Highly-palatable dimension is genetically uncorrelated from the other two, suggesting that independent processes underlie liking high reward foods. This is confirmed by genetic correlations with MRI brain traits which show with distinct associations. Comparison with the corresponding food consumption traits shows a high genetic correlation, while liking exhibits twice the heritability. GWAS analysis identified 1,401 significant food-liking associations which showed substantial agreement in the direction of effects with 11 independent cohorts. In conclusion, we created a comprehensive map of the genetic determinants and associated neurophysiological factors of food-liking.
The ability to taste phenylthiocarbamide (PTC) and 6-n-propylthiouracil (PROP) is a polymorphic trait mediated by the TAS2R38 bitter taste receptor gene. It has long been hypothesized that global ...genetic diversity at this locus evolved under pervasive pressures from balancing natural selection. However, recent high-resolution population genetic studies of TAS2Rs suggest that demographic events have played a critical role in the evolution of these genes. We here utilized the largest TAS2R38 database yet analyzed, consisting of 5,589 individuals from 105 populations, to examine natural selection, haplotype frequencies and linkage disequilibrium to estimate the effects of both selection and demography on contemporary patterns of variation at this locus. We found signs of an ancient balancing selection acting on this gene but no post Out-Of-Africa departures from neutrality, implying that the current observed patterns of variation can be predominantly explained by demographic, rather than selective events. In addition, we found signatures of ancient selective forces acting on different African TAS2R38 haplotypes. Collectively our results provide evidence for a relaxation of recent selective forces acting on this gene and a revised hypothesis for the origins of the present-day worldwide distribution of TAS2R38 haplotypes.
Although several studies demonstrated that platelet count is higher in women, decreases with age, and is influenced by genetic background, most clinical laboratories still use the reference interval ...150-400×10(9) platelets/L for all subjects. The present study was to identify age- and sex-specific reference intervals for platelet count.
We analysed electronic records of subjects enrolled in three population-based studies that investigated inhabitants of seven Italian areas including six geographic isolates. After exclusion of patients with malignancies, liver diseases, or inherited thrombocytopenias, which could affect platelet count, reference intervals were estimated from 40,987 subjects with the non parametric method computing the 2.5° and 97.5° percentiles.
Platelet count was similar in men and women until the age of 14, but subsequently women had steadily more platelets than men. The number of platelets decreases quickly in childhood, stabilizes in adulthood, and further decreases in oldness. The final result of this phenomenon is that platelet count in old age was reduced by 35% in men and by 25% in women compared with early infancy. Based on these findings, we estimated reference intervals for platelet count ×10(9)/L in children (176-452), adult men (141-362), adult women (156-405), old men (122-350) and, old women (140-379). Moreover, we calculated an "extended" reference interval that takes into account the differences in platelet count observed in different geographic areas.
The age-, sex-, and origin-related variability of platelet count is very wide, and the patient-adapted reference intervals we propose change the thresholds for diagnosing both thrombocytopenia and thrombocytosis in Italy.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Genetic variation plays a crucial role in individual differences in food preferences which ultimately influence food selection and health. Our current understanding of this pathway has been informed ...through twin studies (to assess the heritability of food preferences), candidate gene studies, and genome-wide association studies (GWAS). However, most of this literature is mainly focused on genes previously identified as having taste or smell functions. New data suggests that genes not associated with taste or smell perception may be involved in food preferences and contribute to health outcomes. This review highlights these emerging findings and suggests a polygenic risk assessment approach to explore new relationships between food preferences and health risks.
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