Proteolytic cleavage and inactivation of the TRMT1 tRNA modification enzyme by SARS-CoV-2 main protease

Abstract

Nonstructural protein 5 (Nsp5) is the main protease of SARS-CoV-2 that cleaves viral polyproteins into individual polypeptides necessary for viral replication. Here, we show that Nsp5 binds and cleaves human tRNA methyltransferase 1 (TRMT1), a host enzyme required for a common post-transcriptional modification in tRNAs. Human cells infected with SARS-CoV-2 exhibit a decrease in TRMT1 protein levels and TRMT1-catalyzed tRNA modifications, consistent with TRMT1 cleavage and inactivation by Nsp5. Nsp5 cleaves TRMT1 at a specific position that matches the consensus sequence of SARS-CoV-2 polyprotein cleavage sites, and a single mutation within the sequence inhibits Nsp5-dependent proteolysis of TRMT1. The TRMT1 cleavage fragments exhibit altered RNA binding activity and are unable to rescue tRNA modification in TRMT1-deficient human cells. Compared to wildtype human cells, TRMT1-deficient human cells infected with SARS-CoV-2 exhibit reduced levels of intracellular viral RNA. These findings suggest that Nsp5-dependent cleavage of TRMT1 and perturbation of tRNA modification patterns contribute to the cellular pathogenesis and biology of SARS-CoV-2 infection.

Recognition and Cleavage of Human tRNA Methyltransferase TRMT1 by the SARS-CoV-2 Main Protease

“…Our results show that the human tRNAmodifying enzyme TRMT1 is a viable substrate for the SARS-CoV-2 main protease and provide the structural basis for understanding TRMT1 recognition and proteolysis. Importantly, and concurrent with our own work, Zhang et al report that human TRMT1 is cleaved during SARS-CoV-2 infection and that this leads to loss of m2,2G modification in virally infected cells (36). Together, our studies corroborate and characterize cleavage of human TRMT1 by SARS-CoV-2 M pro from the cellular to the atomic level, and raise important new questions about the roles of tRNA methylation during viral infection.…”

“…Cleavage of TRMT1 during viral infection would be predicted to affect host and viral translation and increase cellular sensitivity to redox stress, which could have important implications for understanding the ability of SARS-CoV-2 to hijack host protein synthesis and impact cellular phenotypes. Concurrent with submission of this manuscript, a preprint reporting the observation of M pro -catalyzed TRMT1 cleavage and downregulation of m2,2G modification in SARS-CoV-2-infected human cells was posted, 59 validating our in vitro observations and increasing the importance and relevance of our detailed biochemical and structural data in the context of human viral infection. Additionally, our TRMT1-M pro structure highlights a distinct substrate binding mode that reveals the S3′ pocket on the surface of the main protease.…”

“…Our work defines the structural basis for understanding recognition and cleavage of the human tRNA methyltransferase TRMT1 by the SARS-CoV-2 main protease. Zhang et al provide evidence that TRMT1 is proteolyzed and inactivated during SARS-CoV-2 infection, which would be predicted to affect host and viral translation and increase cellular sensitivity to redox stress (36). TRMT1 cleavage may therefore have important implications for understanding the ability of SARS-CoV-2 to hijack host protein synthesis and impact cellular phenotypes and COVID-19 pathogenesis.…”

SARS-CoV-2 main protease Nsp5 cleaves and inactivates human tRNA methyltransferase TRMT1

“…According to the primary sequence analysis and structure prediction by Alphafold, TRMT1 36–530 contains the S-adenosylmethionine (SAM) binding site, and the cleaved C-terminal portion is probably involved in tRNA binding. During the preparation and submission of this manuscript, two independent studies have been released on bioRxiv ( Oliviera et al., 2023 ; Zhang et al., 2023 ), both revealing the cleavage of TRMT1 by SARS-CoV-2 Nsp5 at Q530, which are in full agreement with our results. Very early after the virus enters human cells, the virus genome generates several non-structural proteins, among which Nsp1 has been widely reported to inhibit mRNA translation by blocking the mRNA entry channel ( Tidu et al., 2020 ).…”

The hidden RNA code: implications of the RNA epitranscriptome in the context of viral infections

“…This suggests a potential correlation between phage codon bias and tRNA modification content ( Lampi et al, 2023 ). On the other hand, TRMT1 and its modifications were reduced in response to SARS-CoV-2 infection, likely due to the activity of specific viral proteases ( D’Oliviera et al, 2023 ; Zhang K. et al, 2023 ) ( Figure 1B ). Interestingly, a multiplex small RNA sequencing analysis of nasopharyngeal swabs showed significant variation of tRNA modification patterns among patients with distinct clinical manifestations of COVID-19 ( Katanski et al, 2022 ).…”