Pervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs

ABSTRACT Pervasive transcription is observed in a wide range of organisms, including humans, mice, and viruses, but the functional significance of the resulting transcripts remains uncertain. Current genetic approaches are often limited by their emphasis on protein-coding open reading frames (ORFs)....

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Autores principales:	Susan P. Canny, Tiffany A. Reese, L. Steven Johnson, Xin Zhang, Amal Kambal, Erning Duan, Catherine Y. Liu, Herbert W. Virgin
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2014
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/9846cf0dfaeb49e396edc879289d299b
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spelling	oai:doaj.org-article:9846cf0dfaeb49e396edc879289d299b2021-11-15T15:45:13ZPervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs10.1128/mBio.01033-132150-7511https://doaj.org/article/9846cf0dfaeb49e396edc879289d299b2014-05-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01033-13https://doaj.org/toc/2150-7511ABSTRACT Pervasive transcription is observed in a wide range of organisms, including humans, mice, and viruses, but the functional significance of the resulting transcripts remains uncertain. Current genetic approaches are often limited by their emphasis on protein-coding open reading frames (ORFs). We previously identified extensive pervasive transcription from the murine gammaherpesvirus 68 (MHV68) genome outside known ORFs and antisense to known genes (termed expressed genomic regions [EGRs]). Similar antisense transcripts have been identified in many other herpesviruses, including Kaposi’s sarcoma-associated herpesvirus and human and murine cytomegalovirus. Despite their prevalence, whether these RNAs have any functional importance in the viral life cycle is unknown, and one interpretation is that these are merely “noise” generated by functionally unimportant transcriptional events. To determine whether pervasive transcription of a herpesvirus genome generates RNA molecules that are functionally important, we used a strand-specific functional approach to target transcripts from thirteen EGRs in MHV68. We found that targeting transcripts from six EGRs reduced viral protein expression, proving that pervasive transcription can generate functionally important RNAs. We characterized transcripts emanating from EGRs 26 and 27 in detail using several methods, including RNA sequencing, and identified several novel polyadenylated transcripts that were enriched in the nuclei of infected cells. These data provide the first evidence of the functional importance of regions of pervasive transcription emanating from MHV68 EGRs. Therefore, studies utilizing mutation of a herpesvirus genome must account for possible effects on RNAs generated by pervasive transcription. IMPORTANCE The fact that pervasive transcription produces functionally important RNAs has profound implications for design and interpretation of genetic studies in herpesviruses, since such studies often involve mutating both strands of the genome. This is a common potential problem; for example, a conservative estimate is that there are an additional 73,000 nucleotides transcribed antisense to annotated ORFs from the 119,450-bp MHV68 genome. Recognizing the importance of considering the function of each strand of the viral genome independently, we used strand-specific approaches to identify six regions of the genome encoding transcripts that promoted viral protein expression. For two of these regions, we mapped novel transcripts and determined that targeting transcripts from these regions reduced viral replication and the expression of other viral genes. This is the first description of a function for these RNAs and suggests that novel transcripts emanating from regions of pervasive transcription are critical for the viral life cycle.Susan P. CannyTiffany A. ReeseL. Steven JohnsonXin ZhangAmal KambalErning DuanCatherine Y. LiuHerbert W. VirginAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 5, Iss 2 (2014)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Susan P. Canny Tiffany A. Reese L. Steven Johnson Xin Zhang Amal Kambal Erning Duan Catherine Y. Liu Herbert W. Virgin Pervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs
description	ABSTRACT Pervasive transcription is observed in a wide range of organisms, including humans, mice, and viruses, but the functional significance of the resulting transcripts remains uncertain. Current genetic approaches are often limited by their emphasis on protein-coding open reading frames (ORFs). We previously identified extensive pervasive transcription from the murine gammaherpesvirus 68 (MHV68) genome outside known ORFs and antisense to known genes (termed expressed genomic regions [EGRs]). Similar antisense transcripts have been identified in many other herpesviruses, including Kaposi’s sarcoma-associated herpesvirus and human and murine cytomegalovirus. Despite their prevalence, whether these RNAs have any functional importance in the viral life cycle is unknown, and one interpretation is that these are merely “noise” generated by functionally unimportant transcriptional events. To determine whether pervasive transcription of a herpesvirus genome generates RNA molecules that are functionally important, we used a strand-specific functional approach to target transcripts from thirteen EGRs in MHV68. We found that targeting transcripts from six EGRs reduced viral protein expression, proving that pervasive transcription can generate functionally important RNAs. We characterized transcripts emanating from EGRs 26 and 27 in detail using several methods, including RNA sequencing, and identified several novel polyadenylated transcripts that were enriched in the nuclei of infected cells. These data provide the first evidence of the functional importance of regions of pervasive transcription emanating from MHV68 EGRs. Therefore, studies utilizing mutation of a herpesvirus genome must account for possible effects on RNAs generated by pervasive transcription. IMPORTANCE The fact that pervasive transcription produces functionally important RNAs has profound implications for design and interpretation of genetic studies in herpesviruses, since such studies often involve mutating both strands of the genome. This is a common potential problem; for example, a conservative estimate is that there are an additional 73,000 nucleotides transcribed antisense to annotated ORFs from the 119,450-bp MHV68 genome. Recognizing the importance of considering the function of each strand of the viral genome independently, we used strand-specific approaches to identify six regions of the genome encoding transcripts that promoted viral protein expression. For two of these regions, we mapped novel transcripts and determined that targeting transcripts from these regions reduced viral replication and the expression of other viral genes. This is the first description of a function for these RNAs and suggests that novel transcripts emanating from regions of pervasive transcription are critical for the viral life cycle.
format	article
author	Susan P. Canny Tiffany A. Reese L. Steven Johnson Xin Zhang Amal Kambal Erning Duan Catherine Y. Liu Herbert W. Virgin
author_facet	Susan P. Canny Tiffany A. Reese L. Steven Johnson Xin Zhang Amal Kambal Erning Duan Catherine Y. Liu Herbert W. Virgin
author_sort	Susan P. Canny
title	Pervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs
title_short	Pervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs
title_full	Pervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs
title_fullStr	Pervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs
title_full_unstemmed	Pervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs
title_sort	pervasive transcription of a herpesvirus genome generates functionally important rnas
publisher	American Society for Microbiology
publishDate	2014
url	https://doaj.org/article/9846cf0dfaeb49e396edc879289d299b
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Pervasive Transcription of a Herpesvirus Genome Generates Functionally Important RNAs

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