Kaposi's Sarcoma-Associated Herpesvirus ORF57 Protein Binds and Protects a Nuclear Noncoding RNA from Cellular RNA Decay Pathways

Brooke B Sahin,Denish Patel,Nicholas K Conrad,Jae U Jung

doi:10.1371/journal.ppat.1000799

Brooke B Sahin, Denish Patel + Show 2 more

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https://doi.org/10.1371/journal.ppat.1000799

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Abstract

The control of RNA stability is a key determinant in cellular gene expression. The stability of any transcript is modulated through the activity of cis- or trans-acting regulatory factors as well as cellular quality control systems that ensure the integrity of a transcript. As a result, invading viral pathogens must be able to subvert cellular RNA decay pathways capable of destroying viral transcripts. Here we report that the Kaposi's sarcoma-associated herpesvirus (KSHV) ORF57 protein binds to a unique KSHV polyadenylated nuclear RNA, called PAN RNA, and protects it from degradation by cellular factors. ORF57 increases PAN RNA levels and its effects are greatest on unstable alleles of PAN RNA. Kinetic analysis of transcription pulse assays shows that ORF57 protects PAN RNA from a rapid cellular RNA decay process, but ORF57 has little effect on transcription or PAN RNA localization based on chromatin immunoprecipitation and in situ hybridization experiments, respectively. Using a UV cross-linking technique, we further demonstrate that ORF57 binds PAN RNA directly in living cells and we show that binding correlates with function. In addition, we define an ORF57-responsive element (ORE) that is necessary for ORF57 binding to PAN RNA and sufficient to confer ORF57-response to a heterologous intronless β-globin mRNA, but not its spliced counterparts. We conclude that ORF57 binds to viral transcripts in the nucleus and protects them from a cellular RNA decay pathway. We propose that KSHV ORF57 protein functions to enhance the nuclear stability of intronless viral transcripts by protecting them from a cellular RNA quality control pathway.

Highlights

Post-transcriptional events in mRNA biogenesis are of central importance to the fidelity and regulation of gene expression
Our published findings using an unusual Kaposi’s sarcoma-associated herpesvirus (KSHV) nuclear RNA, called polyadenylated nuclear (PAN) RNA, have suggested that intronless polyadenylated transcripts are subject to such a quality control system
Because most KSHV genes lack introns, we hypothesized that KSHV must have evolved mechanisms that bypass this quality control system

Summary

Introduction

Post-transcriptional events in mRNA biogenesis are of central importance to the fidelity and regulation of gene expression. Cellular factors regulate nearly every step of RNA metabolism including transcription elongation, RNA splicing, 39 end formation, nuclear export, translation, etc. RNA half-life can be modulated directly, through the activities of regulatory stabilizing or destabilizing protein factors or small RNAs [3,4,5]. RNA quality control pathways ensure aberrant transcripts are less stable than their functional counterparts [6]. Given the importance of these pathways for gene expression, it is no surprise that viruses have evolved mechanisms to counteract pathways that otherwise would lead to the destruction of viral transcripts [3,7]

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