m6A Regulates the Stability of Cellular Transcripts Required for Efficient KSHV Lytic Replication

Manners, Oliver; Baquero-Pérez, Belinda; Mottram, Timothy J.; Yonchev, Ivaylo D; Trevelyan, Christopher J.; Harper, Katherine L.; Menezes, Sarah Bomfim; Patterson, Molly R.; Macdonald, Andrew; Wilson, Stuart A.; Aspden, Julie L.; Whitehouse, Adrian

doi:10.3390/v15061381

Cited by 5 publications

(2 citation statements)

References 74 publications

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“…RNA modifications are central features in post-transcriptional regulation and are known to be dynamic upon viral infection 43 , 53 , 56 . Similarly, previous works have reported the presence of m 6 A modifications in distinct single-stranded cytoplasmic-replicating RNA viruses 38 .…”

Section: Discussionmentioning

confidence: 99%

N6-methyladenosine modification is not a general trait of viral RNA genomes

Baquero-Pérez,

Yonchev,

Delgado-Tejedor

et al. 2024

Nat Commun

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Despite the nuclear localization of the m6A machinery, the genomes of multiple exclusively-cytoplasmic RNA viruses, such as chikungunya (CHIKV) and dengue (DENV), are reported to be extensively m6A-modified. However, these findings are mostly based on m6A-Seq, an antibody-dependent technique with a high rate of false positives. Here, we address the presence of m6A in CHIKV and DENV RNAs. For this, we combine m6A-Seq and the antibody-independent SELECT and nanopore direct RNA sequencing techniques with functional, molecular, and mutagenesis studies. Following this comprehensive analysis, we find no evidence of m6A modification in CHIKV or DENV transcripts. Furthermore, depletion of key components of the host m6A machinery does not affect CHIKV or DENV infection. Moreover, CHIKV or DENV infection has no effect on the m6A machinery’s localization. Our results challenge the prevailing notion that m6A modification is a general feature of cytoplasmic RNA viruses and underscore the importance of validating RNA modifications with orthogonal approaches.

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Section: Discussionmentioning

confidence: 99%

N6-methyladenosine modification is not a general trait of viral RNA genomes

Baquero-Pérez,

Yonchev,

Delgado-Tejedor

et al. 2024

Nat Commun

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“…The cells were incubated for 48 hrs, then harvested and RNA was isolated from the cell pellets. Purified RNA was reverse transcribed, and samples were analysed by qPCR for the levels of ORF57 to determine infectious virion production 33 .…”

Section: Viral Reinfection Assaysmentioning

confidence: 99%

EMG1 methyltransferase activity regulates ribosome occupancy at viral uORFs

Harrington,

Murphy,

Aspden

et al. 2024

Preprint

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Viruses do not encode their own translational machinery and rely exclusively on the translational machinery of the host cell for the synthesis of viral proteins. Viruses have evolved diverse mechanisms to redirect the cellular translation apparatus to favour viral transcripts. A unique mechanism employed by Kaposis sarcoma associated herpesvirus involves manipulation of cellular ribosome composition, producing virus induced specialised ribosomes. These ribosomes scan through KSHV uORFs in late lytic genes, allowing efficient translation of downstream main KSHV ORFs. Herein, we highlight the enhanced association of the ribosomal biogenesis factor EMG1 with precursor 40S ribosome complexes during KSHV lytic replication. Depletion of EMG1 results in significantly reduced expression of viral proteins and progression through the lytic cascade, culminating in a dramatic reduction of infectious virus production. We further demonstrate that the methyltransferase activity of EMG1 is required for effective regulation of translation of KSHV uORFs in late lytic genes.

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Virus-modified paraspeckle-like condensates are hubs for viral RNA processing and their formation drives genomic instability

Harper,

Harrington,

Hayward

et al. 2024

Nat Commun

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The nucleus is a highly organised yet dynamic environment containing distinct membraneless nuclear bodies. This spatial separation enables a subset of components to be concentrated within biomolecular condensates, allowing efficient and discrete processes to occur which regulate cellular function. One such nuclear body, paraspeckles, are comprised of multiple paraspeckle proteins (PSPs) built around the architectural RNA, NEAT1_2. Paraspeckle function is yet to be fully elucidated but has been implicated in a variety of developmental and disease scenarios. We demonstrate that Kaposi’s sarcoma-associated herpesvirus (KSHV) drives formation of structurally distinct paraspeckles with a dramatically increased size and altered protein composition that are required for productive lytic replication. We highlight these virus-modified paraspeckles form adjacent to virus replication centres, potentially functioning as RNA processing hubs for viral transcripts during infection. Notably, we reveal that PSP sequestration into virus-modified paraspeckles result in increased genome instability during both KSHV and Epstein Barr virus (EBV) infection, implicating their formation in virus-mediated tumourigenesis.

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m6A Regulates the Stability of Cellular Transcripts Required for Efficient KSHV Lytic Replication

Cited by 5 publications

References 74 publications

N6-methyladenosine modification is not a general trait of viral RNA genomes

N6-methyladenosine modification is not a general trait of viral RNA genomes

EMG1 methyltransferase activity regulates ribosome occupancy at viral uORFs

Virus-modified paraspeckle-like condensates are hubs for viral RNA processing and their formation drives genomic instability

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