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bioRxiv · 10.1101/2021.04.02.438257

The m6A landscape of polyadenylated nuclear (PAN) RNA and its related methylome in the context of KSHV replication.

Martin, S. E.·Gan, H.·Toomer, G.·Sridhar, N.·Sztuba Solinska, J.
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Abstract

Polyadenylated nuclear (PAN) RNA is a non-coding transcript involved in Kaposis sarcoma-associated herpesvirus (KSHV) lytic reactivation and regulation of cellular and viral gene expression. We have shown that PAN RNA has a dynamic secondary structure and protein binding profiles that can be influenced by the epitranscriptomic modifications. N6-methyladenosine (m6A) is an abundant signature found in viral and virus-encoded RNAs. Here, we combined an antibody-independent next-generation mapping with direct RNA sequencing to elucidate the m6A landscape of PAN RNA during the KSHV latent and lytic stages of infection. Using a newly developed method, termed Selenium-modified deoxythymidine triphosphate reverse transcription and Ligation Assisted PCR analysis of m6A (SLAP), we gained insight into the fraction of modification at identified sites. Using comprehensive proteomic approaches, we identified writers, erasers, and readers that regulate the m6A status of PAN. We verified the temporal and spatial subcellular availability of the methylome components for PAN modification by performing confocal microscopy analysis. Additionally, the RNA biochemical probing outlined structural alterations invoked by m6A in the context of full-length PAN RNA. This work represents the first comprehensive overview of the dynamic interplay between the cellular epitranscriptomic machinery and a specific viral RNA.

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Martin, S. E., Gan, H., Toomer, G., Sridhar, N., Sztuba Solinska, J.. 2021-04-02. The m6A landscape of polyadenylated nuclear (PAN) RNA and its related methylome in the context of KSHV replication.. https://doi.org/10.1101/2021.04.02.438257

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