The interaction of animal cytoplasmic RNA viruses with the nucleus to facilitate replication

Abstract

A number of positive and negative strand RNA viruses whose primary site of replication is the cytoplasm use the nucleus and/or nuclear components in order to facilitate their replicative processes and alter host cell function. The nucleus itself is divided into a number of different sub-domains including structures such as the nucleolus. Many of the nuclear proteins that localise to these domains are involved in RNA processing, and because of their limited coding capacity, it may be necessary for RNA viruses to sequester such cellular factors in order to facilitate the replication, transcription and translation of their genomes. Amongst the best-studied examples of this are the picornaviruses, whose infection results in the redistribution of nuclear proteins to the cytoplasm and their interaction with the internal ribosome entry site (IRES) to facilitate translation of the picornavirus polyprotein. Examples can be found of other positive and also negative strand RNA virus proteins that localise to the nucleus and sub-domains (especially the nucleolus) during virus infection, and several localisation motifs have been defined. Apart from sequestering nuclear proteins for a role in replication, such viruses may also target the nucleus to disrupt nuclear functions and to inhibit antiviral responses.

Fig. 1

In this confocal microscope section, transcription sites are shown green and nuclear speckles are shown red. Overlap between the two compartments (yellow) shows that many transcription sites at the borders of nuclear speckles. The most intense transcription factories are in nucleoli and are remote from speckles. Histone-tagged with green fluorescent protein shows the distribution of DNA. Image courtesy of Dean Jackson (UMIST) and Francisco Iborra (University of Oxford).

Fig. 2

(a) Detection of a nucleolin-GFP fusion protein (green) by indirect immunoflourescence and nuclear DNA (red) by direct fluorescence using a confocal microscope. Rat cardiac myocytes were transfected with a plasmid expressing a nucleolin-GFP fusion protein under the control of a PolII promoter. Nuclear DNA and regions of rRNA transcription (nucleoli) were visualised by staining cells with propidium iodide. White arrows indicate nucleoli. (b) Detection of Cajal bodies (green) by indirect immunoflourescence and nuclear DNA (red) by direct fluorescence using a confocal microscope. Cajal bodies were detected using a rabbit polyclonal anti p80 (coillin) antibody (kindly provided by Professor Angus Lamond).

Fig. 3

Comparison of the intercellular localisation of the avian coronavirus infectious bronchitis virus nucleoprotein (N protein) that localises both to the cytoplasm and nucleolus (a) and a mutant protein that lacks a nucleolar localisation signal (b), which had been identified by sequence comparison to known nucleolar localisation signals (NuLS) (Hiscox et al., 2001). Vero cells were transfected with either a plasmid, pTriExIBVN that expressed a wild-type N protein fused to a C-terminal his-tag (Wurm et al., 2001) (a) or a plasmid, pTriExIBVNΔ_348–372, in which the putative NuLS was deleted by overlapping PCR, no nucleolar localisation is observed (b). IBV N protein (red) and the his-tag (green) were detected by appropriate antibodies. Co-localisation, where it occurs, is yellow. Examples of cells in which N protein has localised to the nucleolus are arrowed. Magnification ×160.