Rat cytomegalovirus major immediate-early enhancer switching results in altered growth characteristics

Area of Science:

• Virology
• Molecular Biology
• Immunology

Background:

• Cytomegalovirus (CMV) tropism and latency are hypothesized to be influenced by the major immediate-early (MIE) enhancer.
• Rat cytomegalovirus (RCMV) and murine CMV (MCMV) show species-specific replication in vivo but differ in cell culture growth.
• Previous studies indicated MCMV can replicate in rat cells, while RCMV cannot grow in murine cells, with tropism not due to entry barriers.

Purpose of the Study:

• To investigate the role of the RCMV MIE enhancer in determining viral tropism and replication.
• To construct a recombinant RCMV with an MCMV MIE enhancer to assess its impact on in vitro and in vivo tropism.
• To understand the contribution of the MIE enhancer to CMV dissemination and latency.

Main Methods:

• Construction of a recombinant RCMV by replacing the native MIE enhancer with the MCMV MIE enhancer.
• Assessment of recombinant virus growth in rat embryo fibroblast (REF) and murine fibroblast cell lines.
• Analysis of MIE transcription, replication kinetics, and tissue tropism in vivo (spleen, salivary gland) following infection.
• Evaluation of viral reactivation from spleen explants post-infection.

Main Results:

• Recombinant RCMV growth in tissue culture remained restricted to rat cells, indicating other factors influence in vitro tropism.
• Despite comparable or increased MIE transcription, the recombinant virus showed decreased replication at low multiplicity of infection.
• In vivo, the recombinant virus replicated normally in the spleen during acute infection but was deficient in spreading to the salivary gland.
• Reactivation from spleen explants was observed four months post-infection, even with the foreign MIE enhancer.

Conclusions:

• The MIE enhancer plays a significant role in RCMV tropism for specific tissues, such as the salivary gland, impacting virus dissemination.
• In vitro tropism of RCMV is not solely determined by the MIE enhancer, suggesting the involvement of other viral or host factors.
• The MIE enhancer's function in CMV tropism extends beyond initiating early gene expression, influencing tissue-specific spread and potentially latency.