Targeting macrophage- and intestinal epithelial cell-specific microRNAs against norovirus restricts replication in vivo.

Lucy Thorne,Dalan Bailey,Jia Lu,Yasmin Chaudhry,Ian Goodfellow

doi:10.1099/jgv.0.001065

Abstract

Until recently, our understanding of the cellular tropism of human norovirus (HuNoV), a major cause of viral gastroenteritis, has been limited. Immune cells and intestinal epithelial cells (IECs) have been proposed as targets of HuNoV replication in vivo, although the contribution of each to pathogenesis and transmission is unknown. Murine norovirus (MNV) is widely used as a surrogate model for HuNoV, as it replicates in cultured immune cells. The importance of the complete MNV immune cell tropism in vivo has not been determined. Recent work has linked replication in IECs to viral persistence in vivo. MNV provides a model to assess the relative contribution of each cell tropism to viral replication in immunocompetent native hosts. Here we exploited cell-specific microRNAs to control MNV replication, through insertion of microRNA target sequences into the MNV genome. We demonstrated the utility of this approach for MNV in vitro by selectively reducing replication in microglial cells, using microglial-specific miR-467c. We then showed that inserting a target sequence for the haematopoietic-specific miR-142-3p abrogated replication in a macrophage cell line. The presence of a target sequence for either miR-142-3p or IEC miR-215 significantly reduced viral secretion during the early stages of a persistent infection in immunocompetent mice, confirming that both cell types support viral replication in vivo. This study provides additional evidence that MNV shares the IEC tropism of HuNoVs in vivo, and now provides a model to dissect the contribution of replication in each cell type to viral pathogenesis and transmission in a native host.

Highlights

Human noroviruses (HuNoVs) are a major cause of viral gastroenteritis worldwide, across all age groups [1], and cause between 2–6 million infections each year in the UK alone [2]
We have exploited the cell-specific regulatory nature of host miRNA expression to further define the cellular tropism of Murine norovirus (MNV) during persistent infection of immunocompetent hosts
Replication in microglial cell lines, we employed haematopoietic-specific miR-142–3p and intestinal epithelial cells (IEC) miR-215 to probe the importance of replication in these cell types in vivo

Summary

Introduction

Human noroviruses (HuNoVs) are a major cause of viral gastroenteritis worldwide, across all age groups [1], and cause between 2–6 million infections each year in the UK alone [2]. A major hurdle in the development of a permissive cell culture system was that, until recently, the cellular tropism of HuNoVs in vivo remained unclear. Evidence of HuNoV antigens in immune cells has been provided by numerous animal infection models, including chimpanzees, piglets and immunocompromised mice [12,13,14]. These studies led to the first demonstration of HuNoV replication in vitro, in a cultured B cell line (BJAB) [10].

Methods

Results

Conclusion