Non-synonymous Nucleotide Changes Research Articles

The novel HLA-A*11:416 allele was identified during high-resolution HLA typing.

HLA-A*11:416 has one non-synonymous nucleotide change from A*11:01:01:01 in codon 39.

The novel HLA-A*02:1009 allele was identified in four unrelated bone marrow donors.

HLA-A*02:1009 has one non-synonymous nucleotide change from A*02:01:01:01 at codon 320.

Characterization of the novel HLA-DQB1*05:277 allele, detected in two unrelated bone marrow donors.

HLA-DQB1*05:277 has one non-synonymous nucleotide change from DQB1*05:01:01:03 at codon 224.

Detection of the novel allele, HLA-A*32:165, in a French individual by next-generation sequencing.

The HLA-A*32:165 allele has one non-synonymous nucleotide change from HLA-A*32:01:01 at nucleotide 242 in exon 2.

The novel HLA-DPB1 allele, HLA-DPB1*1344:01, first identified in Korean individuals by next-generation sequencing.

HLA-DPB1*1344:01 has one non-synonymous nucleotide change from DPB1*02:01:02:01 in codon 117.

The novel HLA-C*06:327 allele was identified in three unrelated bone marrow donors.

HLA-C*06:327 has one non-synonymous nucleotide change from C*06:02:01:01 at codon 222.

Quantifying bacterial evolution in the wild: A birthday problem for Campylobacter lineages.

Measuring molecular evolution in bacteria typically requires estimation of the rate at which nucleotide changes accumulate in strains sampled at different times that share a common ancestor. This approach has been useful for dating ecological and evolutionary events that coincide with the emergence of important lineages, such as outbreak strains and obligate human pathogens. However, in multi-host (niche) transmission scenarios, where the pathogen is essentially an opportunistic environmental organism, sampling is often sporadic and rarely reflects the overall population, particularly when concentrated on clinical isolates. This means that approaches that assume recent common ancestry are not applicable. Here we present a new approach to estimate the molecular clock rate in Campylobacter that draws on the popular probability conundrum known as the ‘birthday problem’. Using large genomic datasets and comparative genomic approaches, we use isolate pairs that share recent common ancestry to estimate the rate of nucleotide change for the population. Identifying synonymous and non-synonymous nucleotide changes, both within and outside of recombined regions of the genome, we quantify clock-like diversification to estimate synonymous rates of nucleotide change for the common pathogenic bacteria Campylobacter coli (2.4 x 10−6 s/s/y) and Campylobacter jejuni (3.4 x 10−6 s/s/y). Finally, using estimated total rates of nucleotide change, we infer the number of effective lineages within the sample time frame–analogous to a shared birthday–and assess the rate of turnover of lineages in our sample set over short evolutionary timescales. This provides a generalizable approach to calibrating rates in populations of environmental bacteria and shows that multiple lineages are maintained, implying that large-scale clonal sweeps may take hundreds of years or more in these species.

Identification and futuristic role of novel polymorphism of caprine PrP gene

The Caprine Prion Protein (PrP) gene polymorphism in three different native Indian goat populations of Southern Odisha, namely Ganjam (a registered breed of India), Ghumusari and Raighar was studied. The 876 bp amplified segment of PrP gene contains full length coding sequence of 771 bp. In Ganjam and Ghumusari goats, any difference of nucleotide sequence was not identified. However, the comparison of nucleotide sequences of Raighar goats and goats of other locality revealed a change in nucleotide at five different positions (G190A, G724A, A727T, C775G and C800T) which includes two non-synonymous nucleotide changes. The non-synonymous nucleotide change resulted a change in amino acid at two different positions (Ser234Cys and Lys246Phe) in mature polypeptide which were not reported earlier and therefore, considered as novel. On the basis of these variants of PrP gene phylogenetic tree was constructed which showed that Ganjam and Raighar goats appeared in different clade. Since any occurrence of Scrapie infection in goats of Odisha was not reported, it can be proposed that these changes in amino acid may be responsible as resistance allele.

Novel polymorphism of HSP70 gene affected caprine physiological vital parameters

The synthesis of heat shock protein 70 (HSP70) is temperature-dependent and its response is considered as a cellular thermometer in response to thermal stress. The variation in HSP70 gene expression has been positively correlated with thermo-tolerance. Three different goat populations (Black Bengal, Ganjam and Raighar) which differ in body size, coat color and production performance were assessed for effect of thermal stress at different temperature humidity index (THI). The physiological vital parameters like rectal temperature (RT), skin temperature (ST), heart rate (HR) and respiration rate (RR) which are related to thermal stress susceptibility were recorded. The genetic variation of HSP70 gene in the three goat populations and its effect on physiological vital parameters related to thermal stress was assessed. A novel polymorphism (C241T) in the exonic region of HSP70 gene with significant (p < .05) statistical association with the physiological vital parameters like RT, ST and RR was identified indicating its role in thermo-tolerance. Highlights The HSP70 gene was found to be polymorphic in Indian goats. The non-synonymous nucleotide change (C241T) resulted change in amino acid Arginine 241 Cysteine in mature polypeptide which were not reported earlier. The constructed phylogenetic tree showed that Ganjam and Raighar goats are more close to each other.

Antagonistic pleiotropy conceals molecular adaptations in changing environments.

The importance of positive selection in molecular evolution is debated. Evolution experiments under invariant laboratory conditions typically show a higher rate of nonsynonymous nucleotide change than that of synonymous change, demonstrating prevalent molecular adaptations. Natural evolution inferred from genomic comparisons, however, almost always exhibits the opposite pattern even among closely related conspecifics, indicative of a paucity of positive selection. We hypothesize that this apparent contradiction is at least in part attributable to ubiquitous and frequent environmental changes in nature, causing nonsynonymous mutations beneficial at one time deleterious soon after because of antagonistic pleiotropy and hindering their fixations relative to synonymous mutations despite continued population adaptations. To test this hypothesis, we performed yeast evolution experiments in changing and corresponding constant environments, followed by genome sequencing of the evolving populations. We observed a lower nonsynonymous to synonymous rate ratio in antagonistic changing environments than in the corresponding constant environments, and the population dynamics of mutations supports our hypothesis. These findings and the accompanying population genetic simulations suggest that molecular adaptation is consistently underestimated in nature due to the antagonistic fitness effects of mutations in changing environments.

Characterization of the novel HLA-DPA1 allele, HLA-DPA1*01:27.

HLA-DPA1*01:27 differs from DPA1*01:03:01:01 by a single non-synonymous nucleotide change in codon 15 (ACG > ATG).

Improved inference of site-specific positive selection under a generalized parametric codon model when there are multinucleotide mutations and multiple nonsynonymous rates

BackgroundAn excess of nonsynonymous substitutions, over neutrality, is considered evidence of positive Darwinian selection. Inference for proteins often relies on estimation of the nonsynonymous to synonymous ratio (ω = dN/dS) within a codon model. However, to ease computational difficulties, ω is typically estimated assuming an idealized substitution process where (i) all nonsynonymous substitutions have the same rate (regardless of impact on organism fitness) and (ii) instantaneous double and triple (DT) nucleotide mutations have zero probability (despite evidence that they can occur). It follows that estimates of ω represent an imperfect summary of the intensity of selection, and that tests based on the ω > 1 threshold could be negatively impacted.ResultsWe developed a general-purpose parametric (GPP) modelling framework for codons. This novel approach allows specification of all possible instantaneous codon substitutions, including multiple nonsynonymous rates (MNRs) and instantaneous DT nucleotide changes. Existing codon models are specified as special cases of the GPP model. We use GPP models to implement likelihood ratio tests for ω > 1 that accommodate MNRs and DT mutations. Through both simulation and real data analysis, we find that failure to model MNRs and DT mutations reduces power in some cases and inflates false positives in others. False positives under traditional M2a and M8 models were very sensitive to DT changes. This was exacerbated by the choice of frequency parameterization (GY vs. MG), with rates sometimes > 90% under MG. By including MNRs and DT mutations, accuracy and power was greatly improved under the GPP framework. However, we also find that over-parameterized models can perform less well, and this can contribute to degraded performance of LRTs.ConclusionsWe suggest GPP models should be used alongside traditional codon models. Further, all codon models should be deployed within an experimental design that includes (i) assessing robustness to model assumptions, and (ii) investigation of non-standard behaviour of MLEs. As the goal of every analysis is to avoid false conclusions, more work is needed on model selection methods that consider both the increase in fit engendered by a model parameter and the degree to which that parameter is affected by un-modelled evolutionary processes.

More than 150 novel variants of HLA class I genes detected in German Stem Cell Donor Registry and UCLA International Cell Exchange samples.

High throughput analysis using amplicon-based next-generation sequencing (NGS) of HLA class I genes in samples of registered stem cell donors of the German Stem Cell Donor Registry Düsseldorf revealed 151 novel variants. In addition, four new variants were identified in well-defined samples obtained from the UCLA International Cell Exchange program. New alleles included 37 HLA-A, 57 HLA-B, and 61 HLA-C variant alleles. All variants were confirmed by NGS of HLA-A, HLA-B, and HLA-C genes including the respective 5' and 3' untranslated regions as well as Sanger sequence analysis. Mainly, the variants encompass single nucleotide changes in intronic as well as exonic parts of the genes. We identified intronic variations in 114 new alleles, nonsynonymous nucleotide changes in 25 alleles, synonymous nucleotide changes in nine alleles, and three hybrid alleles. Four alleles carry exonic deletions or insertions resulting in frameshift of peptide translation. Two novel alleles of HLA-C were shown to result in splicing defects of the transcript. Two alleles showed exonic as well as intronic changes. Thirty-four of the new alleles were found in multiple samples.

Complete sequence of the ATP6 and ND3 mitochondrial genes in breast cancer tissue of postmenopausal women with different body mass indexes.

Due to the fact that mitochondrial defects and oxidative stress have been related with obesity and breast cancer is more aggressive in women with obesity, we investigated if postmenopausal Mexican-Mestizo women with breast cancer presented somatic mutations in the sequence of the ATP6 and/or ND3 genes. Twenty one postmenopausal Mexican-Mestizo women with breast cancer who underwent mastectomy or breast conserving surgery were studied. Height and weight were used to calculate body mass index. DNA from tumor tissue samples and blood leukocytes was amplified by polymerase chain reaction and sequenced the ATP6 and ND3 mitochondrial genes. Ages ranged from 46 to 82. According to World Health Organization criteria among the 21 women, 7 had a normal BMI, 7 were overweight and 7 had obesity. In regard to the molecular study, after sequencing the coding region of ATP6 and ND3 genes of the DNA obtained from both leukocytes and tumor tissue, we did not find somatic mutations. All of the changes that we found in both genes were polymorphisms: in ATP6, we identified in ten patients 3 non-synonymous nucleotide changes and in ND3 we observed that six patients presented polymorphisms, three of them were synonymous and two non-synonymous. To our knowledge, this constitutes the first report where the complete sequence of the ATP6 and ND3 genes has been analyzed in postmenopausal Mexican-Mestizo women with breast cancer and diverse BMI. Our results differ with those reported in Caucasian and Asian populations, possibly due to ethnic differences.

Lipid-tuned Zinc Transport Activity of Human ZnT8 Protein Correlates with Risk for Type-2 Diabetes

Zinc is a critical element for insulin storage in the secretory granules of pancreatic beta cells. The islet-specific zinc transporter ZnT8 mediates granular sequestration of zinc ions. A genetic variant of human ZnT8 arising from a single nonsynonymous nucleotide change contributes to increased susceptibility to type-2 diabetes (T2D), but it remains unclear how the high risk variant (Arg-325), which is also a higher frequency (>50%) allele, is correlated with zinc transport activity. Here, we compared the activity of Arg-325 with that of a low risk ZnT8 variant (Trp-325). The Arg-325 variant was found to be more active than the Trp-325 form following induced expression in HEK293 cells. We further examined the functional consequences of changing lipid conditions to mimic the impact of lipid remodeling on ZnT8 activity during insulin granule biogenesis. Purified ZnT8 variants in proteoliposomes exhibited more than 4-fold functional tunability by the anionic phospholipids, lysophosphatidylcholine and cholesterol. Over a broad range of permissive lipid compositions, the Arg-325 variant consistently exhibited accelerated zinc transport kinetics versus the Trp-form. In agreement with the human genetic finding that rare loss-of-function mutations in ZnT8 are associated with reduced T2D risk, our results suggested that the common high risk Arg-325 variant is hyperactive, and thus may be targeted for inhibition to reduce T2D risk in the general populations.

Genetic variants in ATP6 and ND3 mitochondrial genes are not associated with aggressive prostate cancer in Mexican–Mestizo men with overweight or obesity

Mitochondrial defects have been related to obesity and prostate cancer. We investigated if Mexican–Mestizo men presenting this type of cancer, exhibited somatic mutations of ATP6 and/or ND3.Body mass index (BMI) was determined; the degree of prostate cancer aggressiveness was demarcated by the Gleason score. DNA from tumor tissue and from blood leukocytes was amplified by the polymerase chain reaction and ATP6 and ND3 were sequenced. We included 77 men: 20 had normal BMI, 38 were overweight and 19 had obesity; ages ranged from 52 to 83. After sequencing ATP6 and ND3, from DNA obtained from leukocytes and tumor tissue, we did not find any somatic mutations. All changes observed, in both genes, were polymorphisms. In ATP6 we identified, in six patients, two non-synonymous nucleotide changes and in ND3 we observed that twelve patients presented non-synonymous polymorphisms. To our knowledge, this constitutes the first report where the complete sequences of the ATP6 and ND3 have been analyzed in Mexican–Mestizo men with prostate cancer and diverse BMI. Our results differ with those reported in Caucasian populations, possibly due to ethnic differences.

Identification of a novel HLA-C*03 variant, C*03:303, by sequence-based typing in a Dai Chinese individual.

HLA-C*03:303 has one nonsynonymous nucleotide change from HLA-C*03:04:01:01 in codon 194, exon 4 (GTC>GCC).

Identification of a novel HLA-C allele, HLA-C*07:477 in a Drung Chinese individual.

HLA-C*07:477 has one nonsynonymous nucleotide change from HLA-C*07:02:01:01 in codon 176, exon 3 (AAG>GAG).

Structural characterization and mutational assessment of podocin — A novel drug target to nephrotic syndrome — An in silico approach

Non-synonymous single nucleotide changes (nSNC) are coding variants that introduce amino acid changes in their corresponding proteins. They can affect protein function; they are believed to have the largest impact on human health compared with SNCs in other regions of the genome. Such a sequence alteration directly affects their structural stability through conformational changes. Presence of these conformational changes near catalytic site or active site may alter protein function and as a consequence receptor-ligand complex interactions. The present investigation includes assessment of human podocin mutations (G92C, P118L, R138Q, and D160G) on its structure. Podocin is an important glomerular integral membrane protein thought to play a key role in steroid resistant nephrotic syndrome. Podocin has a hairpin like structure with 383 amino acids, it is an integral protein homologous to stomatin, and acts as a molecular link in a stretch-sensitive system. We modeled 3D structure of podocin by means of Modeller and validated via PROCHECK to get a Ramachandran plot (88.5% in most favored region), main chain, side chain, bad contacts, gauche and pooled standard deviation. Further, a protein engineering tool Triton was used to induce mutagenesis corresponding to four variants G92C, P118L, R138Q and D160G in the wild type. Perusal of energies of wild and mutated type of podocin structures confirmed that mutated structures were thermodynamically more stable than wild type and therefore biological events favored synthesis of mutated forms of podocin than wild type. As a conclusive part, two mutations G92C (-8179.272 kJ/mol) and P118L (-8136.685 kJ/mol) are more stable and probable to take place in podocin structure over wild podocin structure (-8105.622 kJ/mol). Though there is lesser difference in mutated and wild type (approximately, 74 and 35 kJ/mol), it may play a crucial role in deciding why mutations are favored and occur at the genetic level.

Attenuating Mutations in nsP1 Reveal Tissue-Specific Mechanisms for Control of Ross River Virus Infection

Ross River virus (RRV) is one of a group of mosquito-transmitted alphaviruses that cause debilitating, and often chronic, musculoskeletal disease in humans. Previously, we reported that replacement of the nonstructural protein 1 (nsP1) gene of the mouse-virulent RRV strain T48 with that from the mouse-avirulent strain DC5692 generated a virus that was attenuated in a mouse model of disease. Here we find that the six nsP1 nonsynonymous nucleotide differences between strains T48 and DC5692 are determinants of RRV virulence, and we identify two nonsynonymous nucleotide changes as sufficient for the attenuated phenotype. RRV T48 carrying the six nonsynonymous DC5692 nucleotide differences (RRV-T48-nsP1(6M)) was attenuated in both wild-type and Rag1(-/-) mice. Despite the attenuated phenotype, RRV T48 and RRV-T48-nsP1(6M) loads in tissues of wild-type and Rag1(-/-) mice were indistinguishable from 1 to 3 days postinoculation. RRV-T48-nsP1(6M) loads in skeletal muscle tissue, but not in other tissues, decreased dramatically by 5 days postinoculation in both wild-type and Rag1(-/-) mice, suggesting that the RRV-T48-nsP1(6M) mutant is more sensitive to innate antiviral effectors than RRV T48 in a tissue-specific manner. In vitro, we found that the attenuating mutations in nsP1 conferred enhanced sensitivity to type I interferon. In agreement with these findings, RRV T48 and RRV-T48-nsP1(6M) loads were similar in mice deficient in the type I interferon receptor. Our findings suggest that the type I IFN response controls RRV infection in a tissue-specific manner and that specific amino acid changes in nsP1 are determinants of RRV virulence by regulating the sensitivity of RRV to interferon. Arthritogenic alphaviruses, including Ross River virus (RRV), infect humans and cause debilitating pain and inflammation of the musculoskeletal system. In this study, we identified coding changes in the RRV nsP1 gene that control the virulence of RRV and its sensitivity to the antiviral type I interferon response, a major component of antiviral defense in mammals. Furthermore, our studies revealed that the effects of these attenuating mutations are tissue specific. These findings suggest that these mutations in nsP1 influence the sensitivity of RRV to type I interferon only in specific host tissues. The new knowledge gained from these studies contributes to our understanding of host responses that control alphavirus infection and viral determinants that counteract these responses.