Project description:Viral infections can influence the dynamic changes of the epitranscriptome. However, due to the lack of understanding regarding the impact of PRRSV (Porcine reproductive and respiratory syndrome virus) on m6A modifications in host cells in vitro, we investigated the changes in viral and host mRNA N6-methyladenosine (m6A) modifications during the infection of MARC145 cells by the PRRSV SD16 strain. We found that the expression of major epitranscriptomic proteins changed before and after viral infection, reprogramming both the viral and host epitranscriptomes. Specifically, the expression of METTL3 increased after PRRSV infection. Overexpression of METTL3 promoted PRRSV gene expression and replication, while knockdown of METTL3 suppressed PRRSV gene expression and replication. These findings reveal the dynamic characteristics of the mRNA m6A modification program during PRRSV replication in vitro.
Project description:Bats harbor highly virulent viruses that can infect other mammals, including humans, posing questions about their immune tolerance mechanisms. Bat cells employ multiple strategies to limit virus replication and virus-induced immunopathology, but the coexistence of bats and fatal viruses remains poorly understood. Here, we investigated the antiviral RNA interference (RNAi) pathway in bat cells and discovered that they have an enhanced antiviral RNAi response, producing canonical viral small interfering RNAs (vsiRNAs) upon Sindbis virus (SINV) infection that were missing in human cells. Disruption of Dicer function resulted in increased viral load for three different RNA viruses in bat cells, indicating an interferon-independent antiviral pathway. Furthermore, our findings reveal the simultaneous engagement of Dicer and pattern-recognition receptors (PRRs), such as retinoic acid-inducible gene I (RIG-I), with double-stranded RNA, suggesting that Dicer attenuates the interferon response initiation in bat cells. These insights advance our comprehension of the distinctive strategies bats employ to coexist with viruses.
Project description:The RNA modification N6-methyladenosine (m6A) can modulate mRNA fate and thus affect many biological processes. We analyzed m6A modification across the transcriptome following infection by dengue virus (DENV), Zika virus (ZIKV), West Nile virus (WNV), and hepatitis C virus (HCV). We found that infection by these viruses in the Flaviviridae family alters m6A modification of specific cellular transcripts, including RIOK3 and CIRBP. During viral infection, the addition of m6A to RIOK3 promotes its translation, while loss of m6A in CIRBP promotes alternative splicing. Importantly, we found that viral activation of innate immune sensing or the endoplasmic reticulum (ER) stress response contributes to the changes in m6A modification in RIOK3 and CIRBP, respectively. Further, several transcripts with infection-altered m6A profiles, including RIOK3 and CIRBP, encode proteins that influence DENV, ZIKV, and HCV infection. Overall, this work reveals that cellular signaling pathways activated during viral infection lead to alterations in m6A modification of host mRNAs to regulate infection.
Project description:N6-methyladenosine (m6A), the most common internal RNA modification in eukaryotic mRNAs, is described to be abundantly present in the genomes of cytoplasmic-replicating RNA viruses. Yet, how the host nuclear m6A writer has access to the viral RNAs in the cytoplasm and what are the associated biological consequences remain unknown. Here, we comprehensively addressed these questions by combining antibody-dependent (m6A-seq) and antibody-independent (SELECT and nanopore direct RNA sequencing) methods on the cytoplasmic-replicating Chikungunya virus (CHIKV) RNA, and found no evidence of m6A modifications. Moreover, depletion of m6A modification machinery components did not affect CHIKV infection, and CHIKV infection did not alter their cellular localization. Consistent with these observations, no m6A modifications were found in the RNA genome of the dengue virus (DENV), another cytoplasmic-replicating virus. Our results challenge the idea that m6A modification is a general trait of cytoplasmic-replicating RNA viruses and stress the need of confirming antibody-dependent detection of m6A modifications with orthogonal antibody-independent methods.
Project description:N6-methyladenosine (m6A), the most common internal RNA modification in eukaryotic mRNAs, is described to be abundantly present in the genomes of cytoplasmic-replicating RNA viruses. Yet, how the host nuclear m6A writer has access to the viral RNAs in the cytoplasm and what are the associated biological consequences remain unknown. Here, we comprehensively addressed these questions by combining antibody-dependent (m6A-seq) and antibody-independent (SELECT and nanopore direct RNA sequencing) methods on the cytoplasmic-replicating Chikungunya virus (CHIKV) RNA, and found no evidence of m6A modifications. Moreover, depletion of m6A modification machinery components did not affect CHIKV infection, and CHIKV infection did not alter their cellular localization. Consistent with these observations, no m6A modifications were found in the RNA genome of the dengue virus (DENV), another cytoplasmic-replicating virus. Our results challenge the idea that m6A modification is a general trait of cytoplasmic-replicating RNA viruses and stress the need of confirming antibody-dependent detection of m6A modifications with orthogonal antibody-independent methods.