<i>In silico</i>identification of conserved<i>cis</i>-acting RNA elements in the SARS-CoV-2 genome

Alhatlani, Bader Y.

doi:10.1101/2020.06.23.167916

Cited by 3 publications

(4 citation statements)

References 32 publications

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“…2B and 2C ). The first is a C > U substitution at position 241 in the 5′UTR, a region of the genome where RNA secondary structure is highly conserved across Coronavirus species ( Madhugiri et al, 2016 ; Rangan et al, 2020 ; Alhatlani, 2020 ). Using RNAfold ( Gruber et al, 2008 ) we found that this C > U transition had no impact on the stem-loop structure established for SARS-CoV ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Prior scans for positive selection within the SARS-CoV-2 genome used elevated ω as the signal of positive selection, which restricts attention to positive selection based on changes in protein function. For coronaviruses this is a notable limitation, given that many aspects of the lifecycle involve RNA function ( Madhugiri et al, 2016 ; Ziv et al, 2020 ; Alhatlani, 2020 ). In addition, the secondary structure of some segments within the RNA genome is well conserved among coronavirus species, which implies a functional role ( Rangan et al, 2020 ; Sanders et al, 2020 ; Huston et al, 2020 ).…”

Section: Discussionmentioning

confidence: 99%

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Positive selection within the genomes of SARS-CoV-2 and other Coronaviruses independent of impact on protein function

Berrío

Gartner

Wray

2020

PeerJ

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Background The emergence of a novel coronavirus (SARS-CoV-2) associated with severe acute respiratory disease (COVID-19) has prompted efforts to understand the genetic basis for its unique characteristics and its jump from non-primate hosts to humans. Tests for positive selection can identify apparently nonrandom patterns of mutation accumulation within genomes, highlighting regions where molecular function may have changed during the origin of a species. Several recent studies of the SARS-CoV-2 genome have identified signals of conservation and positive selection within the gene encoding Spike protein based on the ratio of synonymous to nonsynonymous substitution. Such tests cannot, however, detect changes in the function of RNA molecules. Methods Here we apply a test for branch-specific oversubstitution of mutations within narrow windows of the genome without reference to the genetic code. Results We recapitulate the finding that the gene encoding Spike protein has been a target of both purifying and positive selection. In addition, we find other likely targets of positive selection within the genome of SARS-CoV-2, specifically within the genes encoding Nsp4 and Nsp16. Homology-directed modeling indicates no change in either Nsp4 or Nsp16 protein structure relative to the most recent common ancestor. These SARS-CoV-2-specific mutations may affect molecular processes mediated by the positive or negative RNA molecules, including transcription, translation, RNA stability, and evasion of the host innate immune system. Our results highlight the importance of considering mutations in viral genomes not only from the perspective of their impact on protein structure, but also how they may impact other molecular processes critical to the viral life cycle.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Positive selection within the genomes of SARS-CoV-2 and other Coronaviruses independent of impact on protein function

Berrío

Gartner

Wray

2020

PeerJ

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“…2) as well other relevant frameshifting and replicationrelated structures (17)(18)(19)(20). Conserved elements at the 5′-and 3′-ends have been identified across many members of the coronavirus family and function as cis-acting elements regulating viral replication (21). Specifically, studies on murine and bovine CoVs showed that phylogenetically conserved SLs in the 5′-UTR are capable of long-range RNA-RNA and protein-RNA interactions responsible for optimal viral replication (22,23).…”

Section: Introductionmentioning

confidence: 99%

Amilorides inhibit SARS-CoV-2 replication in vitro by targeting RNA structures

et al. 2021

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“…Conserved elements at the 5' and 3'-ends have been identified across many members of the coronavirus family and function as cis acting elements regulating viral replication. 20 Specifically, studies on murine and bovine coronaviruses showed that phylogenetically conserved stem loops in the 5'-UTR are capable of long-range RNA-RNA and protein-RNA interactions responsible for optimal viral replication. 21,22 More recently, studies aimed at uncovering the pathway that leads to viral protein synthesis via host cell translation machinery revealed that the presence of the full length 5'-UTR of SARS-CoV-2 leads to a five-fold increase in translation of viral proteins.…”

Section: Introductionmentioning

confidence: 99%

Amilorides inhibit SARS-CoV-2 replication in vitro by targeting RNA structures

Zafferani

Haddad

Luo

et al. 2020

Preprint

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The SARS-CoV-2 pandemic, and the likelihood of future coronavirus pandemics, has rendered our understanding of coronavirus biology more essential than ever. Small molecule chemical probes offer to both reveal novel aspects of virus replication and to serve as leads for antiviral therapeutic development. The RNA-biased amiloride scaffold was recently tuned to target a viral RNA structure critical for translation in enterovirus 71, ultimately uncovering a novel mechanism to modulate positive-sense RNA viral translation and replication. Analysis of CoV RNA genomes reveal many conserved RNA structures in the 5’-UTR and proximal region critical for viral translation and replication, including several containing bulge-like secondary structures suitable for small molecule targeting. Following phylogenetic conservation analysis of this region, we screened an amiloride-based small molecule library against a less virulent human coronavirus, OC43, to identify lead ligands. Amilorides inhibited OC43 replication as seen in viral plaque assays. Select amilorides also potently inhibited replication competent SARS-CoV-2 as evident in the decreased levels of cell free virions in cell culture supernatants of treated cells. Reporter screens confirmed the importance of RNA structures in the 5’-end of the viral genome for small molecule activity. Finally, NMR chemical shift perturbation studies of the first six stem loops of the 5’-end revealed specific amiloride interactions with stem loops 4, 5a, and 6, all of which contain bulge like structures and were predicted to be strongly bound by the lead amilorides in retrospective docking studies. Taken together, the use of multiple orthogonal approaches allowed us to identify the first small molecules aimed at targeting RNA structures within the 5’-UTR and proximal region of the CoV genome. These molecules will serve as chemical probes to further understand CoV RNA biology and can pave the way for the development of specific CoV RNA-targeted antivirals.

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In silicoidentification of conservedcis-acting RNA elements in the SARS-CoV-2 genome

Cited by 3 publications

References 32 publications

Positive selection within the genomes of SARS-CoV-2 and other Coronaviruses independent of impact on protein function

Positive selection within the genomes of SARS-CoV-2 and other Coronaviruses independent of impact on protein function

Amilorides inhibit SARS-CoV-2 replication in vitro by targeting RNA structures

Amilorides inhibit SARS-CoV-2 replication in vitro by targeting RNA structures

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