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The immediate-early (IE) proteins of herpes simplex virus (HSV) function on input genomes and affect many aspects of host cell metabolism to ensure the efficient expression and regulation of the remainder of the genome and, subsequently, the production of progeny virions. Due to the many and varied effects of IE proteins on host cell metabolism, their expression is not conducive to normal cell function and viability. This presents a major impediment to the use of HSV as a vector system. In this study, we describe a series of ICP4 mutants that are defective in different subsets of the remaining IE genes. One mutant,d 109, does not express any of the IE proteins and carries a green fluorescent protein (GFP) transgene under the control of the human cytomegalovirus IE promoter (HCMVIEp).d 109 was nontoxic to Vero and human embryonic lung (HEL) cells at all multiplicities of infection tested and was capable of establishing persistent infections in both of these cell types. Paradoxically, the genetic manipulations that were required to eliminate toxicity and allow the genome to persist in cells for long periods of time also dramatically lowered the level of transgene expression. Efficient expression of the HCMVIEp-GFP transgene in the absence of ICP4 was dependent on the ICP0 protein. Ind 109-infected cells, the level of transgene expression was very low in most cells but abundant in a small subpopulation of cells. However, expression of the transgene could be induced in cells containing quiescentd 109 genomes weeks after the initial infection, demonstrating the functionality of the persisting genomes.

Persistence and Expression of the Herpes Simplex Virus Genome in the Absence of Immediate-Early Proteins