Characterization of pUL5, an HCMV protein interacting with the cellular protein IQGAP1
Giulia Anselmi1, Maria Giuliani1, Giacomo Vezzani1
1GSK, via Fiorentina, 1, 53100, Siena, Italy.
Abstract:
Among the Herpesviridae, human cytomegalovirus (HCMV) owns the largest genome and displays a huge coding potential. Here, we characterized the UL5 gene product (pUL5) of the clinical isolate TR strain. The protein was predicted as a 166-amino-acid membrane protein with a theoretical mass of 19 kDa. Recombinant virus expressing pUL5 with a tag allowed the identification of two pUL5 non-glycosylated species of approximately 19 and 9 kDa, expressed with early and late kinetic respectively. Experiments in infection confirmed that the lower molecular weight species was translated from an internal ATG in the UL5 open reading frame. Confocal microscopy analysis showed that pUL5 localized within the assembly compartment, but is not incorporated in the virion, as shown by Western blot on purified viral particles. Finally, pull-down experiments coupled with mass spectrometry analysis identified IQGAP1 as a pUL5 interactor, giving new hints on possible roles of pUL5 during HCMV infection.
Insights
Researchers characterized the human cytomegalovirus (HCMV) UL5 gene product, pUL5. This protein localizes to the assembly compartment and interacts with IQGAP1, suggesting new roles during HCMV infection.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Human cytomegalovirus (HCMV) possesses the largest genome among Herpesviridae, indicating significant coding potential.
- The UL5 gene product (pUL5) is a poorly understood viral protein.
Purpose of the Study:
- To characterize the expression, localization, and interactions of the HCMV pUL5 protein.
- To elucidate potential functions of pUL5 during the viral replication cycle.
Main Methods:
- Expression and purification of tagged recombinant pUL5.
- Western blot analysis to detect pUL5 species and kinetics.
- Confocal microscopy for pUL5 subcellular localization.
- Pull-down assays with mass spectrometry to identify interacting proteins.
Main Results:
- Two non-glycosylated pUL5 species (19 kDa and 9 kDa) were identified, with distinct early and late expression kinetics.
- The lower molecular weight species is translated from an internal ATG codon.
- pUL5 localizes to the viral assembly compartment but is not incorporated into mature virions.
- IQGAP1 was identified as a pUL5-interacting protein.
Conclusions:
- pUL5 exhibits complex expression kinetics and subcellular localization.
- The interaction with IQGAP1 provides insights into pUL5's role in HCMV infection.
- Further research is warranted to fully understand pUL5 function in viral replication.
