Proteomics Analysis of Herpes Simplex Virus Type 1‑Infected Cells Reveals Dynamic Changes of Viral Protein Expression, Ubiquitylation, and Phosphorylation

Herpesviruses
are among the most complex and widespread human viruses
and cause a number of diseases ranging from cold sores to genital
infections and encephalitis. While the composition of viral particles
has been studied, less is known about the expression of the whole
viral proteome in infected cells. Here, we analyzed the proteome of
the prototypical Herpes Simplex Virus type 1 (HSV1) in infected cells
by mass spectrometry. Using a high sensitivity LTQ-Orbitrap, we achieved
a very high level of protein coverage and identified a total of 67
structural and nonstructural viral proteins. We also identified 90
novel phosphorylation sites and 10 novel ubiquitylation sites on different
viral proteins. Ubiquitylation was observed on nine HSV1 proteins.
We identified phosphorylation sites on about half of the detected
viral proteins; many of the highly phosphorylated ones are known to
regulate gene expression. Treatment with inhibitors of DNA replication
induced changes of both viral protein abundance and modifications,
highlighting the interdependence of viral proteins during the life
cycle. Given the importance of expression dynamics, ubiquitylation,
and phosphorylation for protein function, these findings will serve
as important tools for future studies on herpesvirus biology.