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Human cytomegalovirus UL36 protein is dispensable for viral replication in cultured cells
1Howard Hughes Medical Institute, Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544-1014, USA.
Abstract:
Consistent with earlier analyses of human cytomegalovirus UL36 mRNA, we find that the UL36 protein is present throughout infection. In fact, it is delivered to the infected cell as a constituent of the virion. Curiously, much less UL36 protein accumulated in cells infected with the AD169 strain of human cytomegalovirus than in cells infected with the Towne or Toledo strain, and localization of the protein in cells infected with AD169 is strikingly different from that in cell infected with the Towne or Toledo strain. The variation in steady-state level of the proteins results from different stabilities of the proteins. The UL36 proteins from the three viral strains differ by several amino acid substitutions. However, this variability is not responsible for the different half-lives because the AD169 and Towne proteins, which exhibit very different half-lives within infected cells, exhibit the same half-life when introduced into uninfected cells by transfection with expression plasmids. We demonstrate that the UL36 protein is nonessential for growth in cultured cells, and we propose that the ability of the virus to replicate in the absence of UL36 function likely explains the striking strain-specific variation in the half-life and intracellular localization of the protein.
Insights
Human cytomegalovirus UL36 protein levels and localization vary significantly between viral strains due to protein stability, not amino acid differences. The UL36 protein is nonessential for viral replication in cell culture.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Human cytomegalovirus (HCMV) is a significant pathogen.
- The HCMV UL36 protein is a component of the virion and present during infection.
- Strain-specific variations in HCMV proteins can impact viral pathogenesis.
Purpose of the Study:
- To investigate the strain-specific differences in human cytomegalovirus (HCMV) UL36 protein expression and localization.
- To determine the underlying mechanisms responsible for these observed variations.
- To assess the essentiality of the UL36 protein for viral replication.
Main Methods:
- Western blot analysis to quantify UL36 protein levels in infected cells.
- Immunofluorescence microscopy to visualize UL36 protein localization.
- Pulse-chase experiments to assess protein stability.
- Transfection of expression plasmids to evaluate protein half-life independent of viral infection.
Main Results:
- HCMV UL36 protein accumulation and intracellular localization differ markedly between AD169 and Towne/Toledo strains.
- These variations are attributed to differential protein stability, not amino acid sequence differences, as demonstrated by transfection experiments.
- The UL36 protein is nonessential for HCMV replication in cultured cells.
Conclusions:
- HCMV UL36 protein exhibits strain-dependent stability and localization.
- The non-essential nature of UL36 for viral growth in vitro likely permits the evolution of these strain-specific variations.
- Understanding these variations may provide insights into HCMV strain diversity and adaptation.