Unveiling the role of PUS7-mediated pseudouridylation in host protein interactions specific for the SARS-CoV-2 RNA genome

Giambruno, Roberto; Zacco, Elsa; Ugolini, Camilla; Vandelli, Andrea; Mulroney, Logan; D’Onghia, Manfredi; Giuliani, Bianca; Criscuolo, Elena; Castelli, Matteo; Clementi, Nicola; Clementi, Massimo; Mancini, Nicasio; Bonaldi, Tiziana; Gustincich, Stefano; Leonardi, Tommaso; Tartaglia, Gian Gaetano; Nicassio, Francesco

doi:10.1016/j.omtn.2023.102052

Cited by 6 publications

(5 citation statements)

References 68 publications

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“…It has been widely accepted that Ψ-modified RNAs can suppress innate immune responses and may influence the mRNA vaccine design 74 . Although several studies have reported the presence of Ψ in viral RNAs 75,76 , it has not been thoroughly confirmed with the latest sequencing technologies. We therefore applied BACS to various RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), hepatitis C virus (HCV), Zika virus (ZIKV), hepatitis delta virus (HDV), and Sindbis virus (SINV), to study the existence of Ψ in viral transcripts and genomes.…”

Section: Mapping Of ψ In Viral Rnasmentioning

confidence: 97%

“…We therefore applied BACS to various RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), hepatitis C virus (HCV), Zika virus (ZIKV), hepatitis delta virus (HDV), and Sindbis virus (SINV), to study the existence of Ψ in viral transcripts and genomes. We first focused on SARS-CoV-2, since earlier studies reported Ψ in its genomic and subgenomic RNAs by Nanopore sequencing 75,76 . Similar to previous RNA-seq results 77 , the majority of reads were mapped to the positive strand with a unique pattern at the 3'-end corresponding to the subgenomic RNAs (Supplementary Fig.…”

Section: Mapping Of ψ In Viral Rnasmentioning

confidence: 99%

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Absolute quantitative and base-resolution sequencing reveals comprehensive landscape of pseudouridine across the human transcriptome

Xu,

Kong,

Cheng

et al. 2024

Preprint

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Pseudouridine (Ψ) is one of the most abundant modifications in cellular RNA. However, its function remains elusive, mainly due to the lack of highly sensitive and accurate detection methods. To address this challenge, we introduced 2-bromoacrylamide-assisted cyclization sequencing (BACS) for quantitative profiling of Ψ at single-base resolution. Based on novel bromoacrylamide cyclization chemistry, BACS enables a Ψ-to-C transition. Compared to previous methods, BACS allowed the precise identification of Ψ positions, especially in densely modified Ψ regions and consecutive uridine sequences. BACS successfully detected all known Ψ sites in human rRNA and spliceosomal snRNAs and generated the first quantitative Ψ map of human snoRNA and tRNA. Furthermore, BACS simultaneously detected adenosine-to-inosine (A-to-I) editing sites andN1-methyladenosine (m1A). Depletion of three key pseudouridine synthases (PUS) enabled us to elucidate the targets and sequence motifs of TRUB1, PUS7, and PUS1 in HeLa cells. We further applied BACS to Epstein-Barr virus (EBV)-encoded small RNAs (EBERs) and identified a highly abundant Ψ114site in EBER2. Surprisingly, applying BACS to a panel of RNA viruses demonstrated the absence of Ψ in their viral transcripts or genomes, shedding light on differences in pseudouridylation between virus families. We anticipate BACS to serve as a powerful tool to uncover the biological importance of Ψ in future studies.

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Section: Mapping Of ψ In Viral Rnasmentioning

confidence: 97%

Section: Mapping Of ψ In Viral Rnasmentioning

confidence: 99%

Absolute quantitative and base-resolution sequencing reveals comprehensive landscape of pseudouridine across the human transcriptome

Xu,

Kong,

Cheng

et al. 2024

Preprint

View full text Add to dashboard Cite

show abstract

“…In the heart, CUL3 cooperates with adaptors such as KLHL2 and RHOBTB1 to specifically recognize substrates such as WNK3 and PDE5 for ubiquitination and subsequent proteasomal degradation [112]. Interestingly, WNK3 is the host protein that interacts with SARS-CoV-2 RNA [113].…”

Section: Genes/proteins Identified By 1-l Transcription Of C-(aa)n-n ...mentioning

confidence: 99%

“…SARS-CoV-2 Nsp14 mediates the effects of viral infection on the host cell transcriptome, with BCL2L13 showing downregulated expression [99]. WNK3 is a host protein that interacts with SARS-CoV-2 RNA [113]. MiR-223-3p-loaded exosomes from bronchoalveolar lavage fluid promote alveolar macrophage autophagy and reduce acute lung injury by inhibiting the expression of STK39 [116].…”

Section: Genes/proteins Known To Be Related To Covid-19mentioning

confidence: 99%

1-L Transcription of SARS-CoV-2 Spike Protein S1 Subunit

Nahalka

2024

IJMS

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The COVID-19 pandemic prompted rapid research on SARS-CoV-2 pathogenicity. Consequently, new data can be used to advance the molecular understanding of SARS-CoV-2 infection. The present bioinformatics study discusses the “spikeopathy” at the molecular level and focuses on the possible post-transcriptional regulation of the SARS-CoV-2 spike protein S1 subunit in the host cell/tissue. A theoretical protein–RNA recognition code was used to check the compatibility of the SARS-CoV-2 spike protein S1 subunit with mRNAs in the human transcriptome (1-L transcription). The principle for this method is elucidated on the defined RNA binding protein GEMIN5 (gem nuclear organelle-associated protein 5) and RNU2-1 (U2 spliceosomal RNA). Using the method described here, it was shown that 45% of the genes/proteins identified by 1-L transcription of the SARS-CoV-2 spike protein S1 subunit are directly linked to COVID-19, 39% are indirectly linked to COVID-19, and 16% cannot currently be associated with COVID-19. The identified genes/proteins are associated with stroke, diabetes, and cardiac injury.

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“…First, we categorized putative Ψ positions within a known pseudouridine synthase (PUS) motif. For TRUB1, we searched for the motif GUUCN 42 ; for PUS7, we searched for the motif UNUAR 158 .…”

Section: Expression Profiling Of ψ Modification Machinerymentioning

confidence: 99%

Transcriptome-wide detection and characterization of pseudouridine mRNA modifications across diverse human cell lines

McCormick

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Pseudouridine (Ψ) is among the most abundant RNA modification in the human transcriptome and plays crucial roles in modulating mRNA stability, enhancing translation efficiency, and optimizing mRNA structure. The accurate detection and characterization of Ψ on mRNA is hindered by several factors, including the low abundance of mRNA, tissue-specific and cell-to-cell heterogeneity, and technical limitations. Nanopore direct RNA sequencing (DRS) enables full-length transcript sequencing while preserving post-transcriptional modifications in their native context. This technology allows for transcriptome-wide, amplification-free characterization of modifications at single nucleotide resolution. Differential analysis across different cell types can enhance our understanding of how Ψ influences gene regulation. We previously developed Mod-p ID, leveraging the systematic U-to-C basecalling error caused by the presence of Ψ on a native RNA strand to detect and map relative Ψ occupancy across the human transcriptome by comparing to an unmodified transcriptome. This tool can answer fundamental questions regarding the prevalence, conservation, and cell type-specific expression of Ψ across diverse tissue types.In this dissertation, we 1) develop a universal negative control as a community resource to provide a foundation for transcriptome-wide detection and characterization of RNA modifications, and 2) apply this resource for a comprehensive Ψ analysis across diverse human cell types using Mod-p ID.We first generated and characterized a multicellular in vitro transcribed (IVT), unmodified transcriptome (pan-IVT) from six immortalized human cell lines derived from diverse tissue types to provide a comprehensive negative control for public use within the RNA modification field. IVT libraries consist of paired transcripts assembled from canonical nucleotides using poly-A selected RNA. We show that combining multiple cell lines increases transcriptome coverage, helping to enrich data sets used for RNA modification analysis.We then sequenced the native transcriptomes of six human cell lines using DRS and used the Mod-p ID algorithm to catalog the relative positional occupancy of Ψ in each cell line. We show that Ψ sites can be conserved across cell lines, suggesting the presence of Ψ is important for normal cellular function or is cell-type specific, indicating the Ψ may also be consequential for cell-specific function.The transcriptome-wide annotated datasets constructed in this dissertation provide scientists across the RNA modification field with a negative control to conduct their own RNA modification analysis with the pan-IVT and highlight biologically interesting Ψ sites for robust functionalization studies.

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Unveiling the role of PUS7-mediated pseudouridylation in host protein interactions specific for the SARS-CoV-2 RNA genome

Cited by 6 publications

References 68 publications

Absolute quantitative and base-resolution sequencing reveals comprehensive landscape of pseudouridine across the human transcriptome

Absolute quantitative and base-resolution sequencing reveals comprehensive landscape of pseudouridine across the human transcriptome

1-L Transcription of SARS-CoV-2 Spike Protein S1 Subunit

Transcriptome-wide detection and characterization of pseudouridine mRNA modifications across diverse human cell lines

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