Coding Sequence Variants Research Articles

A Novel Pathogenic HSPG2 Mutation in Schwartz-Jampel Syndrome.

Schwartz–Jampel syndrome is a rare autosomal recessive disease caused by mutation in the heparan sulfate proteoglycan 2 (HSPG2) gene. Its cardinal symptoms are skeletal dysplasia and neuromuscular hyperactivity. Herein, we identified a new pathogenic mutation site (NM_005529.6:c.1125C>G; p.Cys375Trp) of HSPG2 leading to Schwartz–Jampel syndrome by whole-exome sequencing. This mutation carried by the asymptomatic parents was previously registered in a single-nucleotide polymorphism database of the National Institutes of Health as a coding sequence variant rs543805444. The pathogenic nature of this missense mutation was demonstrated by in silico pathogenicity assessment, clinical presentations, and cellular function of primary fibroblast derived from patients. Various in silico software applications predicted the mutation to be pathogenic [Sorting Intolerant From Tolerant (SIFT), 0; Polyphen-2, 1; CADD (Combined Annotation Dependent Depletion), 23.7; MutationTaster, 1; DANN (deleterious annotation of genetic variants using neural networks); 0.9]. Needle electromyography revealed extensive complex repetitive discharges and multiple polyphasic motor unit action potentials in axial and limb muscles at rest. Short exercise test for myotonia showed Fournier pattern I. At cellular levels, mutant primary fibroblasts had reduced levels of secreted perlecan and impaired migration ability but normal capability of proliferation. Patients with this mutation showed more neuromuscular instability and relatively mild skeletal abnormality comparing with previously reported cases.

Widespread misregulation of inter-species hybrid transcriptomes due to sex-specific and sex-chromosome regulatory evolution.

When gene regulatory networks diverge between species, their dysfunctional expression in inter-species hybrid individuals can create genetic incompatibilities that generate the developmental defects responsible for intrinsic post-zygotic reproductive isolation. Both cis- and trans-acting regulatory divergence can be hastened by directional selection through adaptation, sexual selection, and inter-sexual conflict, in addition to cryptic evolution under stabilizing selection. Dysfunctional sex-biased gene expression, in particular, may provide an important source of sexually-dimorphic genetic incompatibilities. Here, we characterize and compare male and female/hermaphrodite transcriptome profiles for sibling nematode species Caenorhabditis briggsae and C. nigoni, along with allele-specific expression in their F1 hybrids, to deconvolve features of expression divergence and regulatory dysfunction. Despite evidence of widespread stabilizing selection on gene expression, misexpression of sex-biased genes pervades F1 hybrids of both sexes. This finding implicates greater fragility of male genetic networks to produce dysfunctional organismal phenotypes. Spermatogenesis genes are especially prone to high divergence in both expression and coding sequences, consistent with a "faster male" model for Haldane's rule and elevated sterility of hybrid males. Moreover, underdominant expression pervades male-biased genes compared to female-biased and sex-neutral genes and an excess of cis-trans compensatory regulatory divergence for X-linked genes underscores a "large-X effect" for hybrid male expression dysfunction. Extensive regulatory divergence in sex determination pathway genes likely contributes to demasculinization of XX hybrids. The evolution of genetic incompatibilities due to regulatory versus coding sequence divergence, however, are expected to arise in an uncorrelated fashion. This study identifies important differences between the sexes in how regulatory networks diverge to contribute to sex-biases in how genetic incompatibilities manifest during the speciation process.

A membrane-bound ankyrin repeat protein confers race-specific leaf rust disease resistance in wheat

Plasma membrane-associated and intracellular proteins and protein complexes play a pivotal role in pathogen recognition and disease resistance signaling in plants and animals. The two predominant protein families perceiving plant pathogens are receptor-like kinases and nucleotide binding-leucine-rich repeat receptors (NLR), which often confer race-specific resistance. Leaf rust is one of the most prevalent and most devastating wheat diseases. Here, we clone the race-specific leaf rust resistance gene Lr14a from hexaploid wheat. The cloning of Lr14a is aided by the recently published genome assembly of ArinaLrFor, an Lr14a-containing wheat line. Lr14a encodes a membrane-localized protein containing twelve ankyrin (ANK) repeats and structural similarities to Ca2+-permeable non-selective cation channels. Transcriptome analyses reveal an induction of genes associated with calcium ion binding in the presence of Lr14a. Haplotype analyses indicate that Lr14a-containing chromosome segments were introgressed multiple times into the bread wheat gene pool, but we find no variation in the Lr14a coding sequence itself. Our work demonstrates the involvement of an ANK-transmembrane (TM)-like type of gene family in race-specific disease resistance in wheat. This forms the basis to explore ANK-TM-like genes in disease resistance breeding.

Divergence in Coding Sequence and Expression of Different Functional Categories of Immune Genes between Two Wild Rodent Species.

Differences in immune function between species could be a result of interspecific divergence in coding sequence and/or expression of immune genes. Here, we investigate how the degree of divergence in coding sequence and expression differs between functional categories of immune genes, and if differences between categories occur independently of other factors (expression level, pleiotropy). To this end, we compared spleen transcriptomes of wild-caught yellow-necked mice and bank voles. Immune genes expressed in the spleen were divided into four categories depending on the function of the encoded protein: pattern recognition receptors (PRR); signal transduction proteins; transcription factors; and cyto- and chemokines and their receptors. Genes encoding PRR and cyto-/chemokines had higher sequence divergence than genes encoding signal transduction proteins and transcription factors, even when controlling for potentially confounding factors. Genes encoding PRR also had higher expression divergence than genes encoding signal transduction proteins and transcription factors. There was a positive correlation between expression divergence and coding sequence divergence, in particular for PRR genes. We propose that this is a result of that divergence in PRR coding sequence leads to divergence in PRR expression through positive feedback of PRR ligand binding on PRR expression. When controlling for sequence divergence, expression divergence of PRR genes did not differ from other categories. Taken together, the results indicate that coding sequence divergence of PRR genes is a major cause of differences in immune function between species.

In situ dissecting the evolution of gene duplication with different histone modification patterns based on high-throughput data analysis in Arabidopsis thaliana.

BackgroundGenetic regulation is known to contribute to the divergent expression of duplicate genes; however, little is known about how epigenetic modifications regulate the expression of duplicate genes in plants.MethodsThe histone modification (HM) profile patterns of different modes of gene duplication, including the whole genome duplication, proximal duplication, tandem duplication and transposed duplication were characterized based on ChIP-chip or ChIP-seq datasets. In this study, 10 distinct HM marks including H2Bub, H3K4me1, H3K4me2, H3K4me3, H3K9ac, H3K9me2, H3K27me1, H3K27me3, H3K36me3 and H3K14ac were analyzed. Moreover, the features of gene duplication with different HM patterns were characterized based on 88 RNA-seq datasets of Arabidopsis thaliana.ResultsThis study showed that duplicate genes in Arabidopsis have a more similar HM pattern than single-copy genes in both their promoters and protein-coding regions. The evolution of HM marks is found to be coupled with coding sequence divergence and expression divergence after gene duplication. We found that functionally selective constraints may impose on epigenetic evolution after gene duplication. Furthermore, duplicate genes with distinct functions have more divergence in histone modification compared with the ones with the same function, while higher expression divergence is found with mutations of chromatin modifiers. This study shows the role of epigenetic marks in regulating gene expression and functional divergence after gene duplication in plants based on sequencing data.

Clinical lessons learned in constitutional hypopituitarism from two decades of experience in a large international cohort.

The international GENHYPOPIT network collects phenotypical data and screens genetic causes of non-acquired hypopituitarism. To describe main phenotype patterns and their evolution through life. Patients were screened according to their phenotype for coding sequence variations in 8 genes: HESX1, LHX3, LHX4, PROP1, POU1F1, TBX19, OTX2 and PROKR2. Among 1213 patients (1143 index cases), the age of diagnosis of hypopituitarism was congenital (24%), in childhood (28%), at puberty (32%), in adulthood (7.2%) or not available (8.8%). Noteworthy, pituitary hormonal deficiencies kept on evolving during adulthood in 49 of patients. Growth Hormone deficiency (GHD) affected 85.8% of patients and was often the first diagnosed deficiency. AdrenoCorticoTropic Hormone deficiency rarely preceded GHD, but usually followed it by over 10years. Pituitary Magnetic Resonance Imaging (MRI) abnormalities were common (79.7%), with 39.4% pituitary stalk interruption syndrome (PSIS). The most frequently associated extrapituitary malformations were ophthalmological abnormalities (16.1%). Prevalence of identified mutations was 7.3% of index cases (84/1143) and 29.5% in familial cases (n=146). Genetic analysis in 449 patients without extrapituitary phenotype revealed 36 PROP1, 2 POU1F1 and 17 TBX19 mutations. This large international cohort highlights atypical phenotypic presentation of constitutional hypopituitarism, such as post pubertal presentation or adult progression of hormonal deficiencies. These results justify long-term follow-up, and the need for systematic evaluation of associated abnormalities. Genetic defects were rarely identified, mainly PROP1 mutations in pure endocrine phenotypes.

Role of Genetic Variation in ABC Transporters in Breast Cancer Prognosis and Therapy Response.

Breast cancer is the most common cancer in women in the world. The role of germline genetic variability in ATP-binding cassette (ABC) transporters in cancer chemoresistance and prognosis still needs to be elucidated. We used next-generation sequencing to assess associations of germline variants in coding and regulatory sequences of all human ABC genes with response of the patients to the neoadjuvant cytotoxic chemotherapy and disease-free survival (n = 105). A total of 43 prioritized variants associating with response or survival in the above testing phase were then analyzed by allelic discrimination in the large validation set (n = 802). Variants in ABCA4, ABCA9, ABCA12, ABCB5, ABCC5, ABCC8, ABCC11, and ABCD4 associated with response and variants in ABCA7, ABCA13, ABCC4, and ABCG8 with survival of the patients. No association passed a false discovery rate test, however, the rs17822931 (Gly180Arg) in ABCC11, associating with response, and the synonymous rs17548783 in ABCA13 (survival) have a strong support in the literature and are, thus, interesting for further research. Although replicated associations have not reached robust statistical significance, the role of ABC transporters in breast cancer should not be ruled out. Future research and careful validation of findings will be essential for assessment of genetic variation which was not in the focus of this study, e.g., non-coding sequences, copy numbers, and structural variations together with somatic mutations.

Hybrid Incompatibilities and Transgressive Gene Expression Between Two Closely Related Subspecies of Drosophila.

Genome-wide assays of expression between species and their hybrids have identified genes that become either over- or underexpressed relative to the parental species (i.e., transgressive). Transgressive expression in hybrids is of interest because it highlights possible changes in gene regulation linked to hybrid dysfunction. Previous studies in Drosophila that used long-diverged species pairs with complete or nearly complete isolation (i.e., full sterility and partial inviability of hybrids) and high-levels of genome misregulation have found correlations between expression and coding sequence divergence. The work highlighted the possible effects of directional selection driving sequence divergence and transgressive expression. Whether the same is true for taxa at early stages of divergence that have only achieved partial isolation remains untested. Here, we reanalyze previously published genome expression data and available genome sequence reads from a pair of partially isolated subspecies of Drosophila to compare expression and sequence divergence. We find a significant correlation in rates of expression and sequence evolution, but no support for directional selection driving transgressive expression in hybrids. We find that most transgressive genes in hybrids show no differential expression between parental subspecies and used SNP data to explore the role of stabilizing selection through compensatory mutations. We also examine possible misregulation through cascade effects that could be driven by interacting gene networks or co-option of off-target cis-regulatory elements.

Rare SCN10A variants associated with cardiac conduction system diseases

Abstract Background The genetic bases of cardiac conduction-system disease (CCSD) range from ion channelopathies to mutations in many other genes. Genome-wide association studies have shown common variants in SCN10A influence cardiac conduction. However, it has not yet to be determined whether vulnerability to CCSD is associated with rare coding sequence variation in the SCN10A gene. Purpose We sought to determine the clinical impact of rare variants in SCN10A in patients with CCSD and classified the variants according to the 2015 American College of Medical Genetics and Genomics (ACMG) standards and guidelines. Methods We performed screening for rare variants (minor allele frequency ≤0.001) in SCN10A in CCSD patients with an onset at a young age under 65 or those who had a family history of pacemaker implantation (PMI) (n=40; 18 female; mean age, 41±18 years). We transiently expressed engineered variants in ND 7/23 cells, and conducted whole-cell voltage clamp experiments to clarify the functional properties of the Nav1.8 current. Results We identified nine rare variants in SCN10A in 7 patients. Two patients were carriers of two rare variants in SCN10A and 5 were carriers of one rare variant in SCN10A. Four patients were affected with sinus node dysfunction, 1 were atrioventricular block, and 2 were both dysfunctions. We performed electrophysiological study for 8 of 9 rare variants. It demonstrated that 2 rare variants showed gain-of-function, and 3 rare variants showed loss-of-function. We finally determined 5 likely pathogenic variants in SCN10A in 5 patients (12.5%) according to the ACMG standards and guidelines. All 5 patients underwent a pacemaker implantation at an average age of 43±16. Conclusions These results demonstrate that SCN10A variants play a pivotal role in enhanced susceptibility of CCSD. We suggest the importance for screening SCN10A variants in clinical settings. Funding Acknowledgement Type of funding source: None

Angiotensin-converting enzymes (ACE, ACE2) gene variants and COVID-19 outcome.

The Angiotensin system is implicated in the pathogenesis of COVID-19. First, ACE2 is the cellular receptor for SARS-CoV-2, and expression of the ACE2 gene could regulate the individuaĺs susceptibility to infection. In addition, the balance between ACE1 and ACE2 activity has been implicated in the pathogenesis of respiratory diseases and could play a role in the severity of COVID-19. Functional ACE1/ACE2 gene polymorphisms have been associated with the risk of cardiovascular and pulmonary diseases, and could thus also contribute to the outcome of COVID-19. We studied 204 COVID-19 patients (137 non-severe and 67 severe-ICU cases) and 536 age-matched controls. The ACE1 insertion/deletion and ACE2 rs2285666 polymorphism were determined. Variables frequencies were compared between the groups by logistic regression. We also sequenced the ACE2 coding nucleotides in a group of patients. Severe COVID-19 was associated with hypertension male gender (p<0.001), hypertension (p=0.006), hypercholesterolaemia (p=0.046), and the ACE1-DD genotype (p=0.049). In the multiple logistic regression hypertension (p=0.02, OR=2.26, 95%CI=1.12-4.63) and male gender (p=0.002; OR=3.15, 95%CI=1.56-6.66) remained as independent significant predictors of severity. The ACE2 polymorphism was not associated with the disease outcome. The ACE2 sequencing showed no coding sequence variants that could explain an increased risk of developing COVID-19. In conclusion, an adverse outcome of COVID-19 was associated with male gender, hypertension, hypercholesterolemia and the ACE1 genotype. Our work suggested that the ACE1-I/D might influence COVID-19 severity, but the effect was dependent on the hypertensive status. This result requires further validation in other large cohorts.

Candidate gene expression and coding sequence variants in Warmblood horses with myofibrillar myopathy.

Myofibrillar myopathy (MFM) of unknown aetiology has recently been identified in Warmblood (WB) horses. In humans, 16 genes have been implicated in various MFM-like disorders. To identify variants in 16 MFM candidate genes and compare allele frequencies of all variants between MFM WB and non-MFM WB and coding variants with moderate or severe predicted effects in MFM WB with publicly available data of other breeds. To compare differential gene expression and muscle fibre contractile force between MFM and non-MFM WB. Case-control. 8 MFM WB, 8 non-MFM WB, 33 other WB, 32 Thoroughbreds, 80 Quarter Horses and 77 horses of other breeds in public databases. Variants were called within transcripts of 16 candidate genes using gluteal muscle mRNA sequences aligned to EquCab3.0 and allele frequencies compared by Fisher's exact test among MFM WB, non-MFM WB and public sequences across breeds. Candidate gene differential expression was determined between MFM and non-MFM WB by fitting a negative binomial generalised log-linear model per gene (false discovery rate <0.05). The maximal isometric force/cross-sectional area generated by isolated membrane-permeabilised muscle fibres was determined. None of the 426 variants identified in 16 candidate genes were associated with MFM including 26 missense variants. Breed-specific differences existed in allele frequencies. Candidate gene differential expression and muscle fibre-specific force did not differ between MFM WB (143.1±34.7kPa) and non-MFM WB (140.2±43.7kPa) (P=.8). RNA-seq-only assays transcripts expressed in skeletal muscle. Other possible candidate genes were not evaluated. Evidence for association of variants with a disease is essential because coding sequence variants are common in the equine genome. Variants identified in MFM candidate genes, including two coding variants offered as commercial MFM equine genetic tests, did not associate with the WB MFM phenotype.

1410-P: Genetics of TG/HDL-C Ratio and Its Change over Time in Nondiabetic Subjects

TG/HDL-C ratio (THR) represents a single inherited surrogate predictor of hyperinsulinemia or insulin resistance that is associated with frank development of T2D, premature aging processes and increased mortality. To identify gene variants for THR and its change over time (THRGC), we conducted GWAS among subjects of European ancestry who had complete data from two exams collected ∼7 years apart from the LLFS (n = 3091), a study with familial clustering of exceptional longevity in the U.S. and Denmark. We sought replication in the FamHS (n = 3108, ∼7 years apart between two visits). Subjects with T2D or using medications for dyslipidemia were excluded from this analysis. THRGC was derived using growth curve modeling. Each phenotype was adjusted for age, sex, PCs and familial membership, and then log-transformed to approximate normality. EPACTS with make-kin and emmax options for familial clustering assuming additive genetic effects was used. P&amp;lt;5e-8 was used to declare genome-wide significance. Heritability estimates reached 38% for THR and 46% for THRGC in the LLFS. GWAS identified two significant loci for THR (LPL-rs328, stop gain, coding sequence variant, MAF=0.11, p=8e-9, replication p=4e-9; BUD13-ZNF259-APOA5-rs2072560, intron variant, MAF=0.07, p=2e-8, replication p=8e-14) and one locus for THRGC (LPL-rs78458743, ∼20 kb downstream, MAF=0.1, p=4e-8, replication p=0.01). The LPL gene encodes lipoprotein lipase that hydrolyzes chylomicrons and VLDL-C. Additionally, linkage scan identified a significant (Lods&amp;gt;3) locus on 3q28 (Lods=4.1) for THRGC in the LLFS. Further query of sequence elements under the linkage peak is underway. In conclusion, this first GWAS of THR identified the LPL gene (including a functional variant rs328) and the BUD13-ZNF259-APOA5 gene cluster for THR variation. The new findings highlight the importance of the LPL and APOA5 gene regulation of THR in subjects without diabetes selected for exceptional survival and healthy aging. Disclosure P. An: None. Y. Lu: None. B. Thyagarajan: None. A.L. Kuipers: None. J.A. Anema: None. I. Miljkovic: None. E.W. Daw: None. M. Province: None. Funding National Institute on Aging (U01AG023712, U01AG023744, U01AG023746, U01AG023749, U01AG023755)

Associations of mitochondrial DNA 3777\u20134679 region mutations with maternally inherited essential hypertensive subjects in China

BackgroundNuclear genome or family mitochondrial screening system has become the hot focus of studies into essential hypertension. The role of mitochondrial DNA (mtDNA) in sporadic Chinese patients with hypertension has not been fully understood. The study was to evaluate the associations of mtDNA mutations with maternally inherited essential hypertensive subjects in China.MethodsFrom June 2009 to June 2016, a total of 800 gender-matched Chinese patients with maternally inherited essential hypertension (MIEH) and control group were 1:1 enrolled in this case-control study. Genomic DNA was extracted from each person’s peripheral blood cells. The main mtDNA locations for MIEH were screened with oligodeoxynucleotides 3777-4679 bp, analyzed and compared with the updated consensus Cambridge Sequence. Pathogenic mtDNA mutations were identified from the mitochondrial map.ResultsMIEH subjects presented significantly higher values than those of control group in abdominal circumference (AC), waist circumference (WC), body mass index (BMI), fasting blood glucose (FBG), triglyceride (TG), low-density lipoprotein cholesterol (LDL) and renal function (P < 0.05). MIEH subjects carried more amino acid changes and coding sequence variants (P < 0.01) than control group. The allele frequencies of the eight single nucleotide polymorphisms (SNPs) were significantly different between the two groups, including m.3970 C > T, m.4048G > A, m.4071C > T, m.4086C > T, m. 4164A > G and m.4248 T > C in ND1 gene, and m.4386 T > C and m.4394C > T in tRNAGln gene(P < 0.001). Fifty-five homoplasmic or heteroplasmic mutations were detected in 5 genes: ND1, tRNAIle, tRNAMet, tRNAGln and ND2 gene. The ND1 gene was the main mutation site, where the most mtDNA mutation was m.3970 C > T.ConclusionsThe mtDNA mutations were involved in the process of MIEH. We identified mitochondrial genetic characteristics in MIEH patients in China. The present research serves as a solid foundation for further detailed research on the association between MIEH and mitochondrial dysfunction, and their causal relationship in Chinese and other populations with a similar lifestyle.

Copy number variations of SHANK3 and related sensory profiles in Egyptian children with autism spectrum disorder

BackgroundCurrent estimates indicate that >80 % of children with autism spectrum disorder (ASD) exhibit concomitant sensory processing problems and hyper- or hypo-reactivity to sensory input. These are now included as diagnostic criteria for ASD in the Diagnostic and Statistical Manual of Mental Disorders—Fifth Edition. Chromosomal rearrangements, copy number variations (CNVs), and coding sequence variants involving >100 genes have been identified in patients with ASD. Studying the CNVs of one such gene, SHANK3, and the associated phenotype in patients with ASD could provide insights that will guide future ASD treatments and interventions. ObjectiveTo assess SHANK3 CNVs in children with ASD and investigate their sensory processing patterns using the Short Sensory Profile (SSP). Subjects and methodsForty children with ASD were assessed using the Autism Diagnostic Interview-Revised. SSP was used to evaluate atypical sensory behavior, e.g., hyper- or hypo-reactivity to sensory input or unusual sensory interests. SHANK3 CNVs were assessed in these children using Multiplex Ligation-dependent Probe Amplification. ResultsOf the 40 cases, 77.5 % showed sensory reactivity symptoms. The greatest difference from normality was observed in the under-responsive/seeks sensation domain, followed by the tactile sensitivity domain, whereas hypo-activity (low-energy/weak domain) was closest to normal. The sensory reactivity symptoms were significantly correlated with the severity of ASD. However, only three of the 40 cases had de novo duplications at 22q13.33. The duplications included SHANK3 in two of the cases and only the distal flanking region of SHANK3 in the third case. All three duplication cases also showed symptoms associated with the low-energy/weak domain. ConclusionWe found that children with ASD exhibited sensory processing problems. The SHANK3 copy number gains found demonstrate the gene dosage effect of SHANK3 in ASD pathogenesis. This study adds to the growing understanding of 22q13 duplications that include SHANK3.

The mutational spectrum of Myocilin gene among familial versus sporadic cases of Juvenile onset open angle glaucoma.

Juvenile onset primary open angle glaucoma (JOAG) is a rare disorder associated with high IOP and progressive optic neuropathy in patients diagnosed before the age of 40 years. While in some populations it has primarily an autosomal dominant pattern of inheritance, in others it occurs in a primarily sporadic form. The main aim of the study was to assess the relative prevalence of Myocilin (MYOC) mutations in familial versus sporadic cases of JOAG. We screened 92 unrelated (sporadic) JOAG patients, and 22 affected families (70 affected members and 36 unaffected) for variations in the MYOC gene. We also analyzed the clinical features associated with these variations. Three coding sequence variants were identified as mutations causing JOAG. Four families segregated distinct mutations at Gly367Arg, and two families at Gln337Arg, while only two sporadic JOAG cases harbored MYOC mutations (Gly367Arg and Gln48His). The frequency of MYOC mutations in familial cases (27%) was significantly higher than in sporadic JOAG cases (2%); p = 0.001. A 90% penetrance for the Gly367Arg variant was seen by the age of 40 years in our patients. Characteristic allele signatures, indicative of specific founder effects, were not observed for the Gly367Arg mutation that was looked for in 12 patients among 2 geographically close families, which harbored this mutation. Our data demonstrated that genetic screening for MYOC mutations should be focused toward cases with familial rather than sporadically occurring JOAG.

Whole genome resequencing of the Iranian native dogs and wolves to unravel variome during dog domestication

BackgroundAdvances in genome technology have simplified a new comprehension of the genetic and historical processes crucial to rapid phenotypic evolution under domestication. To get new insight into the genetic basis of the dog domestication process, we conducted whole-genome sequence analysis of three wolves and three dogs from Iran which covers the eastern part of the Fertile Crescent located in Southwest Asia where the independent domestication of most of the plants and animals has been documented and also high haplotype sharing between wolves and dog breeds has been reported.ResultsHigher diversity was found within the wolf genome compared with the dog genome. A total number of 12.45 million SNPs were detected in all individuals (10.45 and 7.82 million SNPs were identified for all the studied wolves and dogs, respectively) and a total number of 3.49 million small Indels were detected in all individuals (3.11 and 2.24 million small Indels were identified for all the studied wolves and dogs, respectively). A total of 10,571 copy number variation regions (CNVRs) were detected across the 6 individual genomes, covering 154.65 Mb, or 6.41%, of the reference genome (canFam3.1). Further analysis showed that the distribution of deleterious variants in the dog genome is higher than the wolf genome. Also, genomic annotation results from intron and intergenic regions showed that the proportion of variations in the wolf genome is higher than that in the dog genome, while the proportion of the coding sequences and 3′-UTR in the dog genome is higher than that in the wolf genome. The genes related to the olfactory and immune systems were enriched in the set of the structural variants (SVs) identified in this work.ConclusionsOur results showed more deleterious mutations and coding sequence variants in the domestic dog genome than those in wolf genome. By providing the first Iranian dog and wolf variome map, our findings contribute to understanding the genetic architecture of the dog domestication.

Control of translation elongation in health and disease.

ABSTRACTRegulation of protein synthesis makes a major contribution to post-transcriptional control pathways. During disease, or under stress, cells initiate processes to reprogramme protein synthesis and thus orchestrate the appropriate cellular response. Recent data show that the elongation stage of protein synthesis is a key regulatory node for translational control in health and disease. There is a complex set of factors that individually affect the overall rate of elongation and, for the most part, these influence either transfer RNA (tRNA)- and eukaryotic elongation factor 1A (eEF1A)-dependent codon decoding, and/or elongation factor 2 (eEF2)-dependent ribosome translocation along the mRNA. Decoding speeds depend on the relative abundance of each tRNA, the cognate:near-cognate tRNA ratios and the degree of tRNA modification, whereas eEF2-dependent ribosome translocation is negatively regulated by phosphorylation on threonine-56 by eEF2 kinase. Additional factors that contribute to the control of the elongation rate include epigenetic modification of the mRNA, coding sequence variation and the expression of eIF5A, which stimulates peptide bond formation between proline residues. Importantly, dysregulation of elongation control is central to disease mechanisms in both tumorigenesis and neurodegeneration, making the individual key steps in this process attractive therapeutic targets. Here, we discuss the relative contribution of individual components of the translational apparatus (e.g. tRNAs, elongation factors and their modifiers) to the overall control of translation elongation and how their dysregulation contributes towards disease processes.

Molecular Analysis of Neuronal Cell Surface Protein gene; Neuroligin 3 in Autism Spectrum Disorder

Introduction: Autism Spectrum Disorder (ASD) is a complex neurodevelopmental disorder that manifests before the age of three. ASD is characterized by a triad of limited or absent verbal communication, a lack of reciprocal social interaction or responsiveness, and restricted, stereotypic patterns of interests and behavior. More than 1 million cases per year are reported in India. It is now believed that ASD is due to complex genetic and environmental interactions. Changes in over 1,000 genes have been reported to be associated with ASD. Many of the associated genes are involved in the development and functions of the brain. The proposed candidate gene for ASD includes the Neuroligin (NLGN). It is a cell adhesion protein on the postsynaptic membrane involved in Synaptogenesis. Objective: To screen the variants in NLGN3 gene associated with Autism Spectrum Disorder. Methods: 1821 mentally ill children from the North Karnataka region of Karnataka, India were screened as per DSM-V Criteria and 108 autistic were included in the study. Genomic DNA was extracted and the candidate gene was amplified using standard PCR condition. The product was sequenced for genetic variation. Result: one 5' UTR variant, one missense variant, and four 3' UTR variants were recorded. The novel Coding sequence variant c.551 T>C transition was observed in 16 autistics (14.81%). This transition changes Valine to Alanine (p.Val184Ala). This may lead to abnormal neuronal cell surface Protein. Conclusion: This was the first study in Karnataka on genetic aspects of autism. Only one coding sequence variant was observed in 14.81% of our population. This variant may lead to potentially abnormal Neuroligin for the etiology of autism in north Karnataka.

MiRNA Genetic Variants Alter Their Secondary Structure and Expression in Patients With RASopathies Syndromes.

RASopathies are a group of rare genetic diseases caused by germline mutations in genes involved in the RAS–mitogen-activated protein kinase (RAS-MAPK) pathway. Whole-exome sequencing (WES) is a powerful approach for identifying new variants in coding and noncoding DNA sequences, including miRNAs. miRNAs are fine-tuning negative regulators of gene expression. The presence of variants in miRNAs could lead to malfunctions of regulation, resulting in diseases. Here, we identified 41 variants in mature miRNAs through WES analysis in five patients with previous clinical diagnosis of RASopathies syndromes. The pathways, biological processes, and diseases that were over-represented among the target genes of the mature miRNAs harboring variants included the RAS, MAPK, RAP1, and PIK3-Akt signaling pathways, neuronal differentiation, neurogenesis and nervous system development, congenital cardiac defects (hypertrophic cardiomyopathy, dilated cardiomyopathy, and arrhythmogenic right ventricular cardiomyopathy), and the phenotypes and syndromes of RASopathies (Noonan syndrome, Legius syndrome, Costello syndrome, Cafe au lait spots multiple, subaortic stenosis, pulmonary valve stenosis, and LEOPARD syndrome). Furthermore, eight selected variants in nine mature miRNAs (hsa-miR-1304, hsa-miR-146a, hsa-miR-196a2, hsa-miR-499a/hsa-miR-499b, hsa-miR-449b, hsa-miR-548l, hsa-miR-575, and hsa-miR-593) may have caused alterations in the secondary structures of miRNA precursor. Selected miRNAs containing variants such as hsa-miR-146a-3p, hsa-miR-196a-3p, hsa-miR-548l, hsa-miR-449b-5p, hsa-miR-575, and hsa-miR499a-3p could regulate classical genes associated with Rasopathies and RAS-MAPK pathways, contributing to modify the expression pattern of miRNAs in patients. RT-qPCR expression analysis revealed four differentially expressed miRNAs that were downregulated: miRNA-146a-3p in P1, P2, P3, P4, and P5, miR-1304-3p in P2, P3, P4, and P5, miR-196a2-3p in P3, and miR-499b-5p in P1. miR-499a-3p was upregulated in P1, P3, and P5. These results indicate that miRNAs show different expression patterns when these variants are present in patients. Therefore, this study characterized the role of miRNAs harboring variants related to RASopathies for the first time and indicated the possible implications of these variants for phenotypes of RASopathies such as congenital cardiac defects and cardio-cerebrovascular diseases. The expression and existence of miRNA variants may be used in the study of biomarkers of the RASopathies.

The role of noncoding mutations in blood cancers.

ABSTRACTThe search for oncogenic mutations in haematological malignancies has largely focused on coding sequence variants. These variants have been critical in understanding these complex cancers in greater detail, ultimately leading to better disease monitoring, subtyping and prognostication. In contrast, the search for oncogenic variants in the noncoding genome has proven to be challenging given the vastness of the search space, the intrinsic difficulty in assessing the impact of variants that do not code for functional proteins, and our still primitive understanding of the function harboured by large parts of the noncoding genome. Recent studies have broken ground on this quest, identifying somatically acquired and recurrent mutations in the noncoding genome that activate the expression of proto-oncogenes. In this Review, we explore some of the best-characterised examples of noncoding mutations in haematological malignancies, and highlight how a significant majority of these variants impinge on gene regulation through the formation of aberrant enhancers and promoters. We delve into the challenges faced by those that embark on a search for noncoding driver mutations, and provide a framework distilled from studies that have successfully identified such variants to overcome some of the most salient hurdles. Finally, we discuss the current therapeutic strategies being explored to target the oncogenic mechanism supported by recurrent noncoding variants. We postulate that the continued discovery and functional characterisation of somatic variants in the noncoding genome will not only advance our understanding of haematological malignancies, but offer novel therapeutic avenues and provide important insights into transcriptional regulation on a broader scale.