A Viral Nuclear Noncoding RNA Binds Re-localized Poly(A) Binding Protein and Is Required for Late KSHV Gene Expression

Sumit Borah,Joan A Steitz,Nicole Darricarrère,Jinjong Myoung,Alicia Darnell,Blossom Damania

doi:10.1371/journal.ppat.1002300

Abstract

During the lytic phase of infection, the gamma herpesvirus Kaposi's Sarcoma-Associated Herpesvirus (KSHV) expresses a highly abundant, 1.1 kb nuclear noncoding RNA of unknown function. We observe that this polyadenylated nuclear (PAN) RNA avidly binds host poly(A)-binding protein C1 (PABPC1), which normally functions in the cytoplasm to bind the poly(A) tails of mRNAs, regulating mRNA stability and translation efficiency. During the lytic phase of KSHV infection, PABPC1 is re-localized to the nucleus as a consequence of expression of the viral shutoff exonuclease (SOX) protein; SOX also mediates the host shutoff effect in which host mRNAs are downregulated while viral mRNAs are selectively expressed. We show that whereas PAN RNA is not required for the host shutoff effect or for PABPC1 re-localization, SOX strongly upregulates the levels of PAN RNA in transient transfection experiments. This upregulation is destroyed by the same SOX mutation that ablates the host shutoff effect and PABPC1 nuclear re-localization or by removal of the poly(A) tail of PAN. In cells induced into the KSHV lytic phase, depletion of PAN RNA using RNase H-targeting antisense oligonucleotides reveals that it is necessary for the production of late viral proteins from mRNAs that are themselves polyadenylated. Our results add to the repertoire of functions ascribed to long noncoding RNAs and suggest a mechanism of action for nuclear noncoding RNAs in gamma herpesvirus infection.

Highlights

Kaposi’s Sarcoma-Associated Herpesvirus (KSHV) is the causative agent of several human cancers and immunoproliferative disorders, including Kaposi’s Sarcoma, Multicentric Castleman’s Disease and Primary Effusion Lymphoma [1,2]
During the lytic phase of infection of Kaposi’s Sarcoma-Associated Herpesvirus (KSHV), a noncoding viral RNA is synthesized that resembles an messenger RNAs (mRNAs) in that it is transcribed by RNA polymerase II, is methyl-G capped at the 59 end, and is polyadenylated at the 39 end; yet this RNA is never exported to the cytoplasm for translation
We present evidence that the function of this abundant, polyadenylated nuclear (PAN) RNA is to bind poly(A) binding protein, which normally binds poly(A) tails of mRNAs in the cytoplasm but is re-localized into the nucleus during lytic KSHV infection

Summary

Introduction

Kaposi’s Sarcoma-Associated Herpesvirus (KSHV) is the causative agent of several human cancers and immunoproliferative disorders, including Kaposi’s Sarcoma, Multicentric Castleman’s Disease and Primary Effusion Lymphoma [1,2]. KSHV infection is characterized by two states: viral latency and lytic growth. Very few viral genes are expressed, reducing the number of viral epitopes available to trigger a host immune response. Given appropriate but incompletely understood stimuli, the virus activates the lytic program of infection. This is characterized by three ordered waves of viral gene expression producing ‘‘immediate early,’’ ‘‘delayed early’’ and ‘‘late’’ proteins, as well as replication of the viral genome. The new genomes are packaged into virions, which are released from the cell for expansive host infection

Methods

Results

Discussion

Conclusion