Author
Christine Mordstein
Other affiliations: Aarhus University, University of Bath
Bio: Christine Mordstein is an academic researcher from University of Edinburgh. The author has contributed to research in topics: Codon usage bias & Gene expression. The author has an hindex of 5, co-authored 10 publications receiving 103 citations. Previous affiliations of Christine Mordstein include Aarhus University & University of Bath.
Topics: Codon usage bias, Gene expression, RNA splicing, Exon, GC-content
Papers
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TL;DR: An evolutionarily informed approach to attenuation is proposed that, unusually, seeks to increase usage of the already most common synonymous codons in SARS-CoV-2 genes.
Abstract: Large-scale re-engineering of synonymous sites is a promising strategy to generate vaccines either through synthesis of attenuated viruses or via codon optimized genes in DNA vaccines. Attenuation typically relies on de-optimisation of codon pairs and maximization of CpG dinucleotide frequencies. So as to formulate evolutionarily-informed attenuation strategies that aim to force nucleotide usage against the direction favoured by selection, here we examine available whole-genome sequences of SARS-CoV-2 to infer patterns of mutation and selection on synonymous sites. Analysis of mutational profiles indicates a strong mutation bias towards U. In turn, analysis of observed synonymous site composition implicates selection against U. Accounting for dinucleotide effects reinforces this conclusion, observed UU content being a quarter of that expected under neutrality. Possible mechanisms of selection against U mutations includes selection for higher expression, for high mRNA stability or lower immunogenicity of viral genes. Consistent with gene-specific selection against CpG dinucleotides, we observe systematic differences of CpG content between SARS-CoV-2 genes. We propose an evolutionarily-informed approach to attenuation that, unusually, seeks to increase usage of the already most common synonymous codons. Comparable analysis of H1N1 and Ebola finds that GC3 deviated from neutral equilibrium is not a universal feature, cautioning against generalization of results.
68 citations
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TL;DR: It is proposed that splicing promotes the nuclear export of AU-rich mRNAs and that codon- and splicing-dependent effects on expression are under evolutionary pressure in the human genome.
Abstract: In the human genome, most genes undergo splicing, and patterns of codon usage are splicing dependent: guanine and cytosine (GC) content is the highest within single-exon genes and within first exons of multi-exon genes. However, the effects of codon usage on gene expression are typically characterized in unspliced model genes. Here, we measured the effects of splicing on expression in a panel of synonymous reporter genes that varied in nucleotide composition. We found that high GC content increased protein yield, mRNA yield, cytoplasmic mRNA localization, and translation of unspliced reporters. Splicing did not affect the expression of GC-rich variants. However, splicing promoted the expression of AT-rich variants by increasing their steady-state protein and mRNA levels, in part through promoting cytoplasmic localization of mRNA. We propose that splicing promotes the nuclear export of AU-rich mRNAs and that codon- and splicing-dependent effects on expression are under evolutionary pressure in the human genome.
49 citations
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TL;DR: A multiplex assay using well-established SARS-CoV-2 targets alongside a human cellular control and a viral spike-in control, which monitor sample quality and nucleic acid extraction efficiency, and demonstrates the feasibility of establishing a robust RT-qPCR assay at approximately 10% of the cost of equivalent commercial assays, which could benefit low-resource environments and make high-volume testing affordable.
Abstract: With the ongoing COVID-19 (Coronavirus Disease 2019) pandemic, caused by the novel coronavirus SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2), there is a need for sensitive, specific, and affordable diagnostic tests to identify infected individuals, not all of whom are symptomatic. The most sensitive test involves the detection of viral RNA using RT-qPCR (quantitative reverse transcription PCR), with many commercial kits now available for this purpose. However, these are expensive, and supply of such kits in sufficient numbers cannot always be guaranteed. We therefore developed a multiplex assay using well-established SARS-CoV-2 targets alongside a human cellular control (RPP30) and a viral spike-in control (Phocine Herpes Virus 1 [PhHV-1]), which monitor sample quality and nucleic acid extraction efficiency, respectively. Here, we establish that this test performs as well as widely used commercial assays, but at substantially reduced cost. Furthermore, we demonstrate >1,000-fold variability in material routinely collected by combined nose and throat swabbing and establish a statistically significant correlation between the detected level of human and SARS-CoV-2 nucleic acids. The inclusion of the human control probe in our assay therefore provides a quantitative measure of sample quality that could help reduce false-negative rates. We demonstrate the feasibility of establishing a robust RT-qPCR assay at approximately 10% of the cost of equivalent commercial assays, which could benefit low-resource environments and make high-volume testing affordable.
29 citations
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TL;DR: In this article, it was shown that the SARS-CoV-2 mutation rate is at least 49-67% higher than would be estimated based on the rate of appearance of variants in sampled genomes.
Abstract: Owing to a lag between a deleterious mutation's appearance and its selective removal, gold-standard methods for mutation rate estimation assume no meaningful loss of mutations between parents and offspring. Indeed, from analysis of closely related lineages, in SARS-CoV-2 the Ka/Ks ratio was previously estimated as 1.008, suggesting no within-host selection. By contrast, we find a higher number of observed SNPs at 4-fold degenerate sites than elsewhere and, allowing for the virus's complex mutational and compositional biases, estimate that the mutation rate is at least 49-67% higher than would be estimated based on the rate of appearance of variants in sampled genomes. Given the high Ka/Ks one might assume that the majority of such intra-host selection is the purging of nonsense mutations. However, we estimate that selection against nonsense mutations accounts for only â¼10% of all the "missing" mutations. Instead, classical protein-level selective filters (against chemically disparate amino acids and those predicted to disrupt protein functionality) account for many missing mutations. It is less obvious why for an intracellular parasite, amino acid cost parameters, notably amino acid decay rate, are also significant. Perhaps most surprisingly, we also find evidence for real time selection against synonymous mutations that move codon usage away from that of humans. We conclude that there is common intra-host selection on SARS-CoV-2 that acts on nonsense, missense and possibly synonymous mutations. This has implications for methods of mutation rate estimation, for determining times to common ancestry and the potential for intra-host evolution including vaccine escape.
27 citations
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TL;DR: A multiplex assay using well-established SARS-CoV-2 targets alongside internal controls that monitor sample quality and nucleic acid extraction efficiency is developed and established that this test performs as well as widely used commercial assays, but at substantially reduced cost.
Abstract: With the ongoing COVID-19 pandemic, caused by the novel coronavirus SARS-CoV-2, there is need for sensitive, specific and affordable diagnostic tests to identify infected individuals, not all of whom are symptomatic. The most sensitive test involves the detection of viral RNA using RT-qPCR, with many commercial kits now available for this purpose. However, these are expensive and supply of such kits in sufficient numbers cannot always be guaranteed. We therefore developed a multiplex assay using well-established SARS-CoV-2 targets alongside internal controls that monitor sample quality and nucleic acid extraction efficiency. Here, we establish that this test performs as well as widely used commercial assays, but at substantially reduced cost. Furthermore, we demonstrate >1,000-fold variability in material routinely collected by nose-and-throat swabbing. The inclusion of a human control probe in our assay provides additional information that could help reduce false negative rates.
17 citations
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01 Jan 2009
TL;DR: In this article, a review outlines the current understanding of miRNA target recognition in animals and discusses the widespread impact of miRNAs on both the expression and evolution of protein-coding genes.
Abstract: MicroRNAs (miRNAs) are endogenous ∼23 nt RNAs that play important gene-regulatory roles in animals and plants by pairing to the mRNAs of protein-coding genes to direct their posttranscriptional repression. This review outlines the current understanding of miRNA target recognition in animals and discusses the widespread impact of miRNAs on both the expression and evolution of protein-coding genes.
646 citations
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TL;DR: It is found that there is no evidence for significantly more transmissible lineages of SARS-CoV-2 due to recurrent mutations, and recurrent mutations currently in circulation appear to be evolutionary neutral and primarily induced by the human immune system via RNA editing, rather than being signatures of adaptation.
Abstract: COVID-19 is caused by the coronavirus SARS-CoV-2, which jumped into the human population in late 2019 from a currently uncharacterised animal reservoir. Due to this recent association with humans, SARS-CoV-2 may not yet be fully adapted to its human host. This has led to speculations that SARS-CoV-2 may be evolving towards higher transmissibility. The most plausible mutations under putative natural selection are those which have emerged repeatedly and independently (homoplasies). Here, we formally test whether any homoplasies observed in SARS-CoV-2 to date are significantly associated with increased viral transmission. To do so, we develop a phylogenetic index to quantify the relative number of descendants in sister clades with and without a specific allele. We apply this index to a curated set of recurrent mutations identified within a dataset of 46,723 SARS-CoV-2 genomes isolated from patients worldwide. We do not identify a single recurrent mutation in this set convincingly associated with increased viral transmission. Instead, recurrent mutations currently in circulation appear to be evolutionary neutral and primarily induced by the human immune system via RNA editing, rather than being signatures of adaptation. At this stage we find no evidence for significantly more transmissible lineages of SARS-CoV-2 due to recurrent mutations.
269 citations
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TL;DR: An algorithm for pooling subsamples based on the geometry of a hypercube that, at low prevalence, accurately identifies individuals infected with SARS-CoV-2 in a small number of tests and few rounds of testing is proposed.
Abstract: Suppressing infections of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) will probably require the rapid identification and isolation of individuals infected with the virus on an ongoing basis. Reverse-transcription polymerase chain reaction (RT-PCR) tests are accurate but costly, which makes the regular testing of every individual expensive. These costs are a challenge for all countries around the world, but particularly for low-to-middle-income countries. Cost reductions can be achieved by pooling (or combining) subsamples and testing them in groups1-7. A balance must be struck between increasing the group size and retaining test sensitivity, as sample dilution increases the likelihood of false-negative test results for individuals with a low viral load in the sampled region at the time of the test8. Similarly, minimizing the number of tests to reduce costs must be balanced against minimizing the time that testing takes, to reduce the spread of the infection. Here we propose an algorithm for pooling subsamples based on the geometry of a hypercube that, at low prevalence, accurately identifies individuals infected with SARS-CoV-2 in a small number of tests and few rounds of testing. We discuss the optimal group size and explain why, given the highly infectious nature of the disease, largely parallel searches are preferred. We report proof-of-concept experiments in which a positive subsample was detected even when diluted 100-fold with negative subsamples (compared with 30-48-fold dilutions described in previous studies9-11). We quantify the loss of sensitivity due to dilution and discuss how it may be mitigated by the frequent re-testing of groups, for example. With the use of these methods, the cost of mass testing could be reduced by a large factor. At low prevalence, the costs decrease in rough proportion to the prevalence. Field trials of our approach are under way in Rwanda and South Africa. The use of group testing on a massive scale to monitor infection rates closely and continually in a population, along with the rapid and effective isolation of people with SARS-CoV-2 infections, provides a promising pathway towards the long-term control of coronavirus disease 2019 (COVID-19).
162 citations
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TL;DR: The weak selection regime that dominates the evolution of most multicellular eukaryotes provides ample material for functional innovation with relatively little adaptation involved.
130 citations