Identification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III

ABSTRACT Deciphering the molecular pathogenesis of virally induced cancers is challenging due, in part, to the heterogeneity of both viral gene expression and host gene expression. Epstein-Barr virus (EBV) is a ubiquitous herpesvirus prevalent in B-cell lymphomas of immune-suppressed individuals. EB...

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Autores principales:	Joshua E. Messinger, Joanne Dai, Lyla J. Stanland, Alexander M. Price, Micah A. Luftig
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	B lymphocyte Epstein-Barr virus gene expression heterogeneity lymphoma Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/ebbf8fb133d6494886f8ec8bc0926aac
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spelling	oai:doaj.org-article:ebbf8fb133d6494886f8ec8bc0926aac2021-11-15T16:22:10ZIdentification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III10.1128/mBio.01006-192150-7511https://doaj.org/article/ebbf8fb133d6494886f8ec8bc0926aac2019-08-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01006-19https://doaj.org/toc/2150-7511ABSTRACT Deciphering the molecular pathogenesis of virally induced cancers is challenging due, in part, to the heterogeneity of both viral gene expression and host gene expression. Epstein-Barr virus (EBV) is a ubiquitous herpesvirus prevalent in B-cell lymphomas of immune-suppressed individuals. EBV infection of primary human B cells leads to their immortalization into lymphoblastoid cell lines (LCLs), serving as a model of these lymphomas. In previous studies, reports from our laboratory have described a temporal model for immortalization with an initial phase characterized by expression of Epstein-Barr nuclear antigens (EBNAs), high levels of c-Myc activity, and hyperproliferation in the absence of the latent membrane proteins (LMPs), called latency IIb. This is followed by the long-term outgrowth of LCLs expressing the EBNAs along with the LMPs, particularly NFκB-activating LMP1, defining latency III. However, LCLs express a broad distribution of LMP1 such that a subset of these cells express LMP1 at levels similar to those seen in latency IIb, making it difficult to distinguish these two latency states. In this study, we performed mRNA sequencing (mRNA-Seq) on early EBV-infected latency IIb cells and latency III LCLs sorted by NFκB activity. We found that latency IIb transcriptomes clustered independently from latency III independently of NFκB. We identified and validated mRNAs defining these latency states. Indeed, we were able to distinguish latency IIb cells from LCLs expressing low levels of LMP1 using multiplex RNA-fluorescence in situ hybridization (RNA-FISH) targeting EBV EBNA2 or LMP1 and human CCR7 or MGST1. This report defines latency IIb as a bona fide latency state independent from latency III and identifies biomarkers for understanding EBV-associated tumor heterogeneity. IMPORTANCE EBV is a ubiquitous pathogen, with >95% of adults harboring a life-long latent infection in memory B cells. In immunocompromised individuals, latent EBV infection can result in lymphoma. The established expression profile of these lymphomas is latency III, which includes expression of all latency genes. However, single-cell analysis of EBV latent gene expression in these lymphomas suggests heterogeneity where most cells express the transcription factor, EBNA2, and only a fraction of the cells express membrane protein LMP1. Our work describes an early phase after infection where the EBNAs are expressed without LMP1, called latency IIb. However, LMP1 levels within latency III vary widely, making these states hard to discriminate. This may have important implications for therapeutic responses. It is crucial to distinguish these states to understand the molecular pathogenesis of these lymphomas. Ultimately, better tools to understand the heterogeneity of these cancers will support more-efficacious therapies in the future.Joshua E. MessingerJoanne DaiLyla J. StanlandAlexander M. PriceMicah A. LuftigAmerican Society for MicrobiologyarticleB lymphocyteEpstein-Barr virusgene expressionheterogeneitylymphomaMicrobiologyQR1-502ENmBio, Vol 10, Iss 4 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	B lymphocyte Epstein-Barr virus gene expression heterogeneity lymphoma Microbiology QR1-502
spellingShingle	B lymphocyte Epstein-Barr virus gene expression heterogeneity lymphoma Microbiology QR1-502 Joshua E. Messinger Joanne Dai Lyla J. Stanland Alexander M. Price Micah A. Luftig Identification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III
description	ABSTRACT Deciphering the molecular pathogenesis of virally induced cancers is challenging due, in part, to the heterogeneity of both viral gene expression and host gene expression. Epstein-Barr virus (EBV) is a ubiquitous herpesvirus prevalent in B-cell lymphomas of immune-suppressed individuals. EBV infection of primary human B cells leads to their immortalization into lymphoblastoid cell lines (LCLs), serving as a model of these lymphomas. In previous studies, reports from our laboratory have described a temporal model for immortalization with an initial phase characterized by expression of Epstein-Barr nuclear antigens (EBNAs), high levels of c-Myc activity, and hyperproliferation in the absence of the latent membrane proteins (LMPs), called latency IIb. This is followed by the long-term outgrowth of LCLs expressing the EBNAs along with the LMPs, particularly NFκB-activating LMP1, defining latency III. However, LCLs express a broad distribution of LMP1 such that a subset of these cells express LMP1 at levels similar to those seen in latency IIb, making it difficult to distinguish these two latency states. In this study, we performed mRNA sequencing (mRNA-Seq) on early EBV-infected latency IIb cells and latency III LCLs sorted by NFκB activity. We found that latency IIb transcriptomes clustered independently from latency III independently of NFκB. We identified and validated mRNAs defining these latency states. Indeed, we were able to distinguish latency IIb cells from LCLs expressing low levels of LMP1 using multiplex RNA-fluorescence in situ hybridization (RNA-FISH) targeting EBV EBNA2 or LMP1 and human CCR7 or MGST1. This report defines latency IIb as a bona fide latency state independent from latency III and identifies biomarkers for understanding EBV-associated tumor heterogeneity. IMPORTANCE EBV is a ubiquitous pathogen, with >95% of adults harboring a life-long latent infection in memory B cells. In immunocompromised individuals, latent EBV infection can result in lymphoma. The established expression profile of these lymphomas is latency III, which includes expression of all latency genes. However, single-cell analysis of EBV latent gene expression in these lymphomas suggests heterogeneity where most cells express the transcription factor, EBNA2, and only a fraction of the cells express membrane protein LMP1. Our work describes an early phase after infection where the EBNAs are expressed without LMP1, called latency IIb. However, LMP1 levels within latency III vary widely, making these states hard to discriminate. This may have important implications for therapeutic responses. It is crucial to distinguish these states to understand the molecular pathogenesis of these lymphomas. Ultimately, better tools to understand the heterogeneity of these cancers will support more-efficacious therapies in the future.
format	article
author	Joshua E. Messinger Joanne Dai Lyla J. Stanland Alexander M. Price Micah A. Luftig
author_facet	Joshua E. Messinger Joanne Dai Lyla J. Stanland Alexander M. Price Micah A. Luftig
author_sort	Joshua E. Messinger
title	Identification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III
title_short	Identification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III
title_full	Identification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III
title_fullStr	Identification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III
title_full_unstemmed	Identification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III
title_sort	identification of host biomarkers of epstein-barr virus latency iib and latency iii
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/ebbf8fb133d6494886f8ec8bc0926aac
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Identification of Host Biomarkers of Epstein-Barr Virus Latency IIb and Latency III

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