Multidimensional analysis of Gammaherpesvirus RNA expression reveals unexpected heterogeneity of gene expression.
PLoS Pathog
; 15(6): e1007849, 2019 06.
Article
em En
| MEDLINE
| ID: mdl-31166996
ABSTRACT
Virus-host interactions are frequently studied in bulk cell populations, obscuring cell-to-cell variation. Here we investigate endogenous herpesvirus gene expression at the single-cell level, combining a sensitive and robust fluorescent in situ hybridization platform with multiparameter flow cytometry, to study the expression of gammaherpesvirus non-coding RNAs (ncRNAs) during lytic replication, latent infection and reactivation in vitro. This method allowed robust detection of viral ncRNAs of murine gammaherpesvirus 68 (γHV68), Kaposi's sarcoma associated herpesvirus and Epstein-Barr virus, revealing variable expression at the single-cell level. By quantifying the inter-relationship of viral ncRNA, viral mRNA, viral protein and host mRNA regulation during γHV68 infection, we find heterogeneous and asynchronous gene expression during latency and reactivation, with reactivation from latency identified by a distinct gene expression profile within rare cells. Further, during lytic replication with γHV68, we find many cells have limited viral gene expression, with only a fraction of cells showing robust gene expression, dynamic RNA localization, and progressive infection. Lytic viral gene expression was enhanced in primary fibroblasts and by conditions associated with enhanced viral replication, with multiple subpopulations of cells present in even highly permissive infection conditions. These findings, powered by single-cell analysis integrated with automated clustering algorithms, suggest inefficient or abortive γHV infection in many cells, and identify substantial heterogeneity in viral gene expression at the single-cell level.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Replicação Viral
/
RNA Mensageiro
/
RNA Viral
/
Regulação Viral da Expressão Gênica
/
Gammaherpesvirinae
/
Infecções por Herpesviridae
/
RNA não Traduzido
Tipo de estudo:
Prognostic_studies
Limite:
Animals
/
Humans
Idioma:
En
Revista:
PLoS Pathog
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
Estados Unidos