1887

Abstract

Cytomegalovirus (CMV) major immediate–early protein 1 (IE1) has multiple functions and is important for efficient viral infection. As does its counterpart in human CMV, murine CMV (MCMV) IE1 also functions as a disruptor of mouse-cell nuclear domain 10 (ND10), where many different gene-regulation proteins congregate. It still remains unclear how MCMV IE1 disperses ND10 and whether this dispersion could have any effect on viral replication. MCMV IE1 is 595 aa long and has multiple functional domains that have not yet been fully analysed. In this study, we dissected the IE1 molecule by truncation and/or deletion and found that the H2B homology domain (amino acid sequence NDIFERI) is required for the dispersion of ND10 by IE1. Furthermore, we made additional deletions and point mutations and found that the minimal truncation in the H2B homology domain required for IE1 to lose the ability to disperse ND10 is just 3 aa (IFE). Surprisingly, the mutated IE1 still interacted with PML and co-localized with ND10 but failed to disperse ND10. This suggests that binding to ND10 key protein is essential to, but not sufficient for, the dispersal of ND10, and that some other unknown mechanism must be involved in this biological procedure. Finally, we generated MCMV with IFE-deleted IE1 (MCMVdlIFE) and its revertant (MCMVIFERQ). Although MCMVdlIFE lost the ability to disperse ND10, plaque assays and viral gene production assays showed that the deletion of IFE did not increase viral replication in cell culture. We conclude that the dispersion of ND10 appears not to be important for MCMV replication in a mouse-cell culture.

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2011-09-01
2019-10-15
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vol. , part 9, pp. 2006–2019

Immunofluorescence assay to determine the physical relationship of IE3 with IE1dlIFE. Immunofluorescence assay to determine the physical relationship of IE3 with PML. Internal primers used for generating mutations by overlapping PCR Molecular cloning, plasmids, and site-directed mutagenesis via BAC system.

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