Cumulative Effects Of Mutations Research Articles

Probing structural basis for enhanced binding of SARS-CoV-2 P.1 variant spike protein with the human ACE2 receptor.

The initial step of infection by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) involves the binding of receptor binding domain (RBD) of the spike protein to the angiotensin converting enzyme 2 (ACE2) receptor. Each successive wave of SARS‐CoV‐2 reports emergence of many new variants, which is associated with mutations in the RBD as well as other parts of the spike protein. These mutations are reported to have enhanced affinity towards the ACE2 receptor as well as are also crucial for the virus transmission. Many computational and experimental studies have demonstrated the effect of individual mutation on the RBD‐ACE2 binding. However, the cumulative effect of mutations on the RBD and away from the RBD was not investigated in detail. We report here a comparative analysis on the structural communication and dynamics of the RBD and truncated S1 domain of spike protein in complex with the ACE2 receptor from SARS‐CoV‐2 wild type and its P.1 variant. Our integrative network and dynamics approaches highlighted a subtle conformational changes in the RBD as well as truncated S1 domain of spike protein at the protein contact level, responsible for the increased affinity with the ACE2 receptor. Moreover, our study also identified the commonalities and differences in the dynamics of the interactions between spike protein of SARS‐CoV‐2 wild type and its P.1 variant with the ACE2 receptor. Further, our investigation yielded an understanding towards identification of the unique RBD residues crucial for the interaction with the ACE2 host receptor. Overall, the study provides an insight for designing better therapeutics against the circulating P.1 variants as well as other future variants.

P–427 Maternal and fetal thrombophilia mutations and the ANXA5 M2 haplotype are more prevalent in first trimester miscarriages of euploid embryos

Abstract Study question Is there a cumulative effect of shared maternal and fetal thrombophilia mutations and the ANXA M2 haplotype in first trimester miscarriages and are there shared genotypes that may predict outcome? Summary answer Coexistence of MTHFR–677CT and PAI–1–4G/5G-mutations in euploid-fetus-mother -pairs may adversely affect early development.Maternal FII–20210GA and PAI–14G mutation were more prevalent in women who miscarried euploid-fetuses. What is known already Spontaneous abortion (SA) is a significant clinical problem with several different etiologies. Certain thrombophilia and folate-related gene mutations have been associated with an increased risk of miscarriage which may be due to impaired coagulation homeostasis. The M2-haplotype of ANXA5-gene promotor encoding the anticoagulation protein Annexin-A5 has also been implicated in early and recurrent SA. However, one important deficiency in the majority of the studies on this subject is a lack of information on the fetal/embryonic contribution to the thrombotic events during placentation and early embryo development. The purpose of this study was to evaluate the prevalence of specific and cumulative thrombophilia mutations in women experiencing first trimester (T1) miscarriages and in their fetal products of conception. Study design, size, duration This is a single Centre retrospective cohort study where we analyzed the frequency and cumulative effect of mutations in 10 thrombophilia/folate related genes and the ANXA5 M2 haplotype in first trimester miscarriages from 229 mother-product of conception (POC) pairs from Jan. 2018-Nov. 2020. Participants/materials, setting, methods All 229 POC DNA samples were confirmed to be fetal and had aneuploidy screening with NGS. We analyzed 10 different mutations in thrombophilia- and folate-related genes (Factor V-Leiden G1691A, Factor V-H1299R, Factor II-G20210A, Factor XIII-V34L, PAI-I–675 4G/5G, FGB–455G/A, MTHFR-C677T and -A1298C, MTR-A2756G, and MTRR- A66G) using single base sequencing methodology and the M2 haplotype of ANXA5 promoter by Sanger sequencing. Maternal–euploid fetus pairs were compared to maternal-aneuploid fetus pairs. SSPS software was used for statistical analyses, and p < 0.05 with CI 95% was considered significant. Main results and the role of chance Aneuploidy was detected in 40.6% (93/229) of POC, and the rest were euploid. Women with euploid fetuses had a significantly higher FII20210GA mutation frequency than those who had aneuploid fetuses (p = 0.02). However, this difference was not detected in the fetuses. Minor allele frequency and genotype frequencies of MTHFR A1298C, MTR A2756G and MTRR A66G mutations as well as for Factor V-Leiden G1691A, Factor V-H1299R and mutations were similar between the groups studied. The frequency of the M2 ANXA5 haplotype was similar between the two groups (3.33% maternal-euploid fetus-pairs vs 3.1% in maternal-aneuploid fetus pairs). PAI-I –675 4G/4G genotype was more frequent in women with euploid fetal losses (25%) vs aneuploid (15%) (p = 0.05), but with no-significant difference in euploid (20%) vs aneuploid fetuses (13%). A cumulative effect of maternal and fetal mutations in MTHFR 677CT and PAI–14G was observed in maternal-euploid fetus pairs (p = 0.0011, OR 3.8 95%CI [1.8–11.3], and p = 0.05, OR 1.2 95%CI [1.12–3.6], respectively). Overall mutation load in tested samples was 4.14 mutations±1.5 [ 1.46±0.8-thrombophilia and 2.45±1.16-folate metabolism] and was similar between both groups of fetuses and their mothers. Limitations, reasons for caution The retrospective nature and cohort design limited the generalizability of our results. The comparison group is maternal-aneuploid fetus- pairs as the reason for the miscarriage is the chromosomal abnormality in the fetus. The ethnic background may be a cofounding factor that needs to be evaluated in a larger study which is ongoing in our center. Wider implications of the findings: Our study found a high mutational load in the 10 folate metabolism and thrombophilia genes we studied in the maternal-fetus pairs. Our findings provide evidence that fetal contribution to placental thrombophilia should be considered in cumulative effect estimations of specific maternal mutations in improvement of management of early miscarriages. Trial registration number Not applicable

A novel case of concurrent occurrence of demyelinating-polyneuropathy-causing PMP22 duplication and SOX10 gene mutation producing severe hypertrophic neuropathy

BackgroundHereditary motor and sensory neuropathy, also referred to as Charcot–Marie–Tooth disease (CMT), is most often caused by a duplication of the peripheral myelin protein 22 (PMP22) gene. This duplication causes CMT type 1A (CMT1A). CMT1A rarely occurs in combination with other hereditary neuromuscular disorders. However, such rare genetic coincidences produce a severe phenotype and have been reported in terms of “double trouble” overlapping syndrome. Waardenburg syndrome (WS) is the most common form of a hereditary syndromic deafness. It is primarily characterized by pigmentation anomalies and classified into four major phenotypes. A mutation in the SRY sex determining region Y-box 10 (SOX10) gene causes WS type 2 or 4 and peripheral demyelinating neuropathy, central dysmyelinating leukodystrophy, WS, and Hirschsprung disease. We describe a 11-year-old boy with extreme hypertrophic neuropathy because of a combination of CMT1A and WS type 2. This is the first published case on the co-occurrence of CMT1A and WS type 2.Case presentationThe 11-year-old boy presented with motor developmental delay and a deterioration in unstable walking at 6 years of age. In addition, he had congenital hearing loss and heterochromia iridis. The neurological examination revealed weakness in the distal limbs with pes cavus. He was diagnosed with CMT1A by the fluorescence in situ hybridization method. His paternal pedigree had a history of CMT1A. However, no family member had congenital hearing loss. His clinical manifestation was apparently severe than those of his relatives with CMT1A. In addition, a whole-body magnetic resonance neurography revealed an extreme enlargement of his systemic cranial and spinal nerves. Subsequently, a genetic analysis revealed a heterozygous frameshift mutation c.876delT (p.F292Lfs*19) in the SOX10 gene. He was eventually diagnosed with WS type 2.ConclusionsWe described a patient with a genetically confirmed overlapping diagnoses of CMT1A and WS type 2. The double trouble with the genes created a significant impact on the peripheral nerves system. Severe phenotype in the proband can be attributed to the cumulative effect of mutations in both PMP22 and SOX10 genes, responsible for demyelinating neuropathy.

Inferring the distribution of fitness effects of spontaneous mutations in Chlamydomonas reinhardtii.

Spontaneous mutations are the source of new genetic variation and are thus central to the evolutionary process. In molecular evolution and quantitative genetics, the nature of genetic variation depends critically on the distribution of effects of mutations on fitness and other quantitative traits. Spontaneous mutation accumulation (MA) experiments have been the principal approach for investigating the overall rate of occurrence and cumulative effect of mutations but have not allowed the phenotypic effects of individual mutations to be studied directly. Here, we crossed MA lines of the green alga Chlamydomonas reinhardtii with its unmutated ancestral strain to create haploid recombinant lines, each carrying an average of 50% of the accumulated mutations in a large number of combinations. With the aid of the genome sequences of the MA lines, we inferred the genotypes of the mutations, assayed their growth rate as a measure of fitness, and inferred the distribution of fitness effects (DFE) using a Bayesian mixture model. We infer that the DFE is highly leptokurtic (L-shaped). Of mutations with absolute fitness effects exceeding 1%, about one-sixth increase fitness in the laboratory environment. The inferred distribution of effects for deleterious mutations is consistent with a strong role for nearly neutral evolution. Specifically, such a distribution predicts that nucleotide variation and genetic variation for quantitative traits will be insensitive to change in the effective population size.

Familial thrombotic risk based on the genetic background of Protein C Deficiency in a Portuguese Study

Inherited protein C (PC) deficiency is a well-known risk factor for venous thrombosis (VT). Plasma PC levels are reliable in moderate to severe deficiencies; however, in mildly deficient individuals, the levels may overlap with those considered normal. Genetic studies of PROC, which encodes PC, could help identify carriers; genome-wide association studies (GWAS) have shown that approximately 50% of phenotypic variation in PC deficiency is caused by the cumulative effects of mutations in several other loci, namely in the PROCR. With the main objective of determining the genotype/phenotype correlation in 59 Portuguese individuals from 26 unrelated families with history of thrombosis and repeatedly low/borderline PC plasma levels, we conducted a molecular study by direct sequencing of PROC; PROC promoter haplotypes and PROCR c.4600A>G polymorphism (rs867186), which are known to influence plasma PC concentrations, were also screened. Twelve different PROC mutations were identified, one of them not previously reported, p.Cys105Arg. The mutation types and locations as well as haplotype combinations correlated with the phenotypic severity. The most frequent mutation, p.Arg199X, correlated with the CGTC haplotype and was identified in nine families containing patients with higher numbers of VT episodes. This mutation in homozygous individuals for the CGTC haplotype is a significant risk factor for VT in Portuguese. These genetic family studies allowed the identification of the unknown carriers and individuals at a higher thrombotic risk within each family, thus permitting the evaluation of the need for prophylactic measures, particularly in at-risk situations.

Mutaciones puntuales en los genes dhfr y dhps de Plasmodium falciparum de tres regiones endémicas

Plasmodium falciparum has the ability to counter the antiparasitic activity of sulphadoxine-pyrimethamine by progressively accumulating mutations in the dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) genes. These mutations gradually increase the resistance of the parasite to these drugs and lead to therapeutic failure. To determine the frequency of mutations associated with resistance to sulphadoxine and pyrimethamine in the dhfr and dhps genes of P. falciparum in samples from patients in three endemic zones of Colombia -La Carpa, Guaviare; Casuarito, Vichada; and Tierralta and Puerto Libertador, Córdoba. Forty samples were selected from patients with uncomplicated P. falciparum malaria. The frequency profiles of the 108, 59 and 164 alleles of dhfr were obtained by application of an allele-specific polymerase chain reaction, whereas the other alleles (alleles 51 of the dhfr gene and 436, 437 and 540 of dhps) were obtained by polymerase chain reaction and restriction fragment length polymorphism. The 108N and 51I mutations in the dhfr gene were found in all of the 40 samples. No mutant alleles were found in the 59 and 164 codons of the dhfr gene, or in the 436 codon of the dhps gene. The 437G mutation was observed in 36 samples and the wild-type allele was present in 3 from Tierralta and one from La Carpa. The 540E mutation was only detected in two samples from Casuarito. The 108N, 51I and 437G mutations prevail in the populations of P. falciparum, indicating a cumulative effect of mutations and the need to continue surveillance for other changes which can lead to the total loss of the efficacy of sulphadoxine-pyrimethamine.

Spontaneous Mutational and Standing Genetic (Co)variation at Dinucleotide Microsatellites in Caenorhabditis briggsae and Caenorhabditis elegans

Understanding the evolutionary processes responsible for shaping genetic variation within and between species requires separating the effects of mutation and selection. Differences between the patterns of genetic variation observed in nature and when mutations are allowed to accumulate in the relative absence of selection can reveal biases imposed by selection. We characterize the genetic variation at dinucleotide microsatellite repeats in four sets of 250-generation mutation accumulation (MA) lines, two in the species Caenorhabditis briggsae and two in Caenorhabditis elegans, and compare the mutational variation with the standing variation in those species. We also compare the mutational properties of microsatellites with the cumulative effects of mutations on fitness in the same lines. Integrated over the whole genome, we infer that the mutation rate of C. briggsae is about twice that of C. elegans, consistent with the cumulative mutational effects on fitness. The mutational spectrum (ratio of insertions to deletions) differs between repeat types and, in some cases, between species. The per-locus mutation rate is significantly positively correlated with the standing genetic variation at the same locus in both species, providing justification for the common practice of using the standing genetic variance as a surrogate for the mutation rate.

Mutational Bias for Body Size in Rhabditid Nematodes

Mutational bias is a potentially important agent of evolution, but it is difficult to disentangle the effects of mutation from those of natural selection. Mutation-accumulation experiments, in which mutations are allowed to accumulate at very small population size, thus minimizing the efficiency of natural selection, are the best way to separate the effects of mutation from those of selection. Body size varies greatly among species of nematode in the family rhabditidae; mutational biases are both a potential cause and a consequence of that variation. We report data on the cumulative effects of mutations that affect body size in three species of rhabditid nematode that vary fivefold in adult size. Results are very consistent with previous studies of mutations underlying fitness in the same strains: two strains of Caenorhabditis briggsae decline in body size about twice as fast as two strains of C. elegans, with a concomitant higher point estimate of the genomic mutation rate; the confamilial Oscheius myriophila is intermediate. There is an overall mutational bias, such that mutations reduce size on average, but the bias appears consistent between species. The genetic correlation between mutations that affect size and those underlying fitness is large and positive, on average.

Role of the Inhibitory Quotient in HIV Therapy

A systemic review is presented of all studies that have evaluated the inhibitory quotient (IQ). The IQ is defined as the ratio between (trough) drug concentration and level of drug resistance of the HIV isolate. From the studies presented, it can be concluded that for protease inhibitors (PIs) and efavirenz, the phenotypic IQ is associated with virological response. The genotypic IQ (GIQ) for PIs was also demonstrated to be associated with virological response. An intrinsic limitation of the GIQ is that it is only applicable for PIs, of which resistance is based on the cumulative effect of mutations. As the IQ can be modified by adjustment of the drug dosage, it may be of clinical value. Its application in patient care should therefore be further investigated.

Point mutations in dihydrofolate reductase and dihydropteroate synthase genes of Plasmodium falciparum isolates from Venezuela.

The present study was designed to characterize mutations in dihydrofolate reductase (DHFR) and dihydropteroate synthase (DHPS) genes of Plasmodium falciparum in the Bolivar region of Venezuela, where high levels of clinical resistance to sulfadoxine-pyrimethamine (SP, Fansidar; F. Hoffman-La Roche, Basel, Switzerland) has been documented. We used a nested mutation-specific polymerase chain reaction and restriction digestion methods to measure 1) the prevalence of DHFR mutations at 16, 50, 51, 59, 108, and 164 codon positions, and 2) the prevalence of mutations in the 436, 437, 581, and 613 codon sites in DHPS gene. In the case of the DHFR gene, of the 54 parasite isolates analyzed, we detected the presence of Asn-108 and Ile-51 in 96% of the isolates and Arg-50 mutation in 64% of the isolates. Each of these mutations has been associated with high level of resistance to pyrimethamine. Only 2 samples (4%) showed the wild type Ser-108 mutation and none showed Thr-108 and Val-16 mutations that are specific for resistance to cycloguanil. In the case of DHPS gene, we found a mutation at position 437 (Gly) in 100% of the isolates and Gly-581 in 96% of the isolates. The simultaneous presence of mutations Asn-108 and Ile-51 in the DHFR gene and Gly-437 and Gly-581 in the DHPS gene in 96% of the samples tested suggested that a cumulative effect of mutations could be the major mechanism conferring high SP resistance in this area.

Cumulative Effects of Mutations in Newly Synthesized Mitochondrial Aspartate Aminotransferase on Uptake into Mitochondria

Mutant genes were constructed which coded for the precursor form of mitochondrial aspartate aminotransferase in which residue cysteine 166 was mutated to either serine or alanine and or forms of the protein lacking both the presequence and residues 1-9 of the mature protein but carrying the same cysteine mutations. The protein products of all of these mutant genes were imported into mitochondria that had been added to the expression system but with varying degrees of efficiency. The results showed that the effects of mutation of cysteine 166 and of deletion of residues 1-9 of the mature protein on sequestration into mitochondria were essentially cumulative, suggesting that these parts of the protein are involved in distinct steps on the recognition/uptake pathway.

Inheritance of low-level resistance to penicillin, tetracycline, and chloramphenicol in Neisseria gonorrhoeae.

The genetics of low-level resistance to penicillin and other antibiotics in a clinical isolate and a multistep laboratory mutant of Neisseria gonorrhoea was studied by transformation. Mutations at three loci affected sensitivity to penicillin. Mutation at penA resulted in an eightfold increase in resistance to penicillin without affecting response to other antimicrobial agents. Mutation at ery resulted in a two- to fourfold increase in resistance to penicillin and similar increases in resistance to many other antibiotics, dyes, and detergents. Mutation at penB resulted in a fourfold increase in resistance to penicillin and tetracycline, the phenotypic expression of which was dependent on the presence of mutation at ery. The cumulative effect of mutations at penA, ery, and penB was an approximate 128-fold increase in penicillin resistance, to a minimum inhibitory concentration of 1.0 mug/ml. Low-level resistance to tetracycline or chloramphenicol was due to similar additive effects between mutations at the nonspecific ery and penB loci and a locus specific for resistance to each drug (tet and chl, respectively). No evidence was found for penicillinases or other drug-inactivating enzymes.