We employed targeted genetic-proteomic approaches to study the virus-host interface either from the virus or host perspective. Using a library of tagged replication competent HCMV and HIV mutant viruses, we infected primary human fibroblasts (for HCMV) and CEM T cells (for HIV), and employed cryogenic cell lysis and rapid immunoaffinity purifications on magnetic beads to isolate virus-host assemblies. For studies on histone deacetylases (HDAC) during viral infections, we generated cell lines stably expressing green fluorescent protein tagged HDACs and probed their interactions and deacetylation activity. Isolated protein complexes were analyzed using a MALDI LTQ Orbitrap (Thermo Fisher Scientific) and the specificity of observed interactions was confirmed by immunofluorescence, reciprocal immunoprecipitation and metabolic labeling with stable isotopes (I-DIRT).
Two interesting findings will be highlighted: 1) studies on pUL32, pUL99, pUL83 and pTRS1 HCMV proteins demonstrated that parallel processes occur at distinct cellular sites during the assembly of HCMV virions, and 2) chromatin remodeling complexes, including histone deacetylases, are targeted by viruses, possibly in part to gain control over host gene expression and modulate the outcome of an infection.
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