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Mol Biol Cell. 2017 Feb 1;28(3):476-487. doi: 10.1091/mbc.E16-08-0612. Epub 2016 Nov 30.

Diverse activities of viral cis-acting RNA regulatory elements revealed using multicolor, long-term, single-cell imaging.

Author information

1
McArdle Laboratory for Cancer Research and Institute for Molecular Virology, University of Wisconsin-Madison, Madison, WI 53706.
2
Morgridge Institute for Research, University of Wisconsin-Madison, Madison, WI 53706.
3
Department of Statistics, University of Wisconsin-Madison, Madison, WI 53706.
4
Laboratory for Optical and Computational Instrumentation and Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53706.
5
McArdle Laboratory for Cancer Research and Institute for Molecular Virology, University of Wisconsin-Madison, Madison, WI 53706 nsherer@wisc.edu ahlquist@wisc.edu.
6
Howard Hughes Medical Institute, University of Wisconsin-Madison, Madison, WI 53706.

Abstract

Cis-acting RNA structural elements govern crucial aspects of viral gene expression. How these structures and other posttranscriptional signals affect RNA trafficking and translation in the context of single cells is poorly understood. Herein we describe a multicolor, long-term (>24 h) imaging strategy for measuring integrated aspects of viral RNA regulatory control in individual cells. We apply this strategy to demonstrate differential mRNA trafficking behaviors governed by RNA elements derived from three retroviruses (HIV-1, murine leukemia virus, and Mason-Pfizer monkey virus), two hepadnaviruses (hepatitis B virus and woodchuck hepatitis virus), and an intron-retaining transcript encoded by the cellular NXF1 gene. Striking behaviors include "burst" RNA nuclear export dynamics regulated by HIV-1's Rev response element and the viral Rev protein; transient aggregations of RNAs into discrete foci at or near the nuclear membrane triggered by multiple elements; and a novel, pulsiform RNA export activity regulated by the hepadnaviral posttranscriptional regulatory element. We incorporate single-cell tracking and a data-mining algorithm into our approach to obtain RNA element-specific, high-resolution gene expression signatures. Together these imaging assays constitute a tractable, systems-based platform for studying otherwise difficult to access spatiotemporal features of viral and cellular gene regulation.

PMID:
27903772
PMCID:
PMC5341730
DOI:
10.1091/mbc.E16-08-0612
[Indexed for MEDLINE]
Free PMC Article

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