title: RNA interference-mediated targeting of human cytomegalovirus immediate-early or early gene products inhibits viral replication with differential effects on cellular functions.
abstract: Viral drug toxicity, resistance, and an increasing immunosuppressed population warrant continued research into new avenues for limiting diseases associated with human cytomegalovirus (HCMV). In this study, a small interfering RNA (siRNA), siX3, was designed to target coding sequences within shared exon 3 of UL123 and UL122 transcripts encoding IE1 and IE2 immediate-early proteins of HCMV. Pretreatment of cells with siX3 reduced the levels of viral protein expression, DNA replication, and progeny virus production compared to control siRNA. Two siRNAs against UL54 and overlapping transcripts (UL55-57) were compared to siX3 in HCMV infection and were also found to be effective at inhibiting HCMV replication. Further investigation into the effects of the siRNAs on viral replication showed that pretreatment with each of the siRNAs resulted in an inhibition in the formation of mature replication compartments. The ability of these siRNAs to prevent or reduce certain cytopathic effects associated with HCMV infection was also examined. Infected cells pretreated with siX3, but not siUL54, retained promyelocytic leukemia (PML) protein in cellular PML bodies, an essential component of this host intrinsic antiviral defense. DNA damage response proteins, which are localized in nuclear viral replication compartments, were reduced in the siX3- and siUL54-treated cells. siX3, but not siUL54, prevented DNA damage response signaling early after infection. Therapeutic efficacy was demonstrated by treating cells with siRNAs after HCMV replication had commenced. Together, these findings suggest that siRNAs targeting exon 3 of the major IE genes or the UL54-57 transcripts be further studied for their potential development into anti-HCMV therapeutics.
authors.affiliation: Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
authors.forename: E
authors.initials: E
authors.lastname: Xiaofei
authors.forename: Bradford M
authors.initials: BM
authors.lastname: Stadler
authors.forename: Michelle
authors.initials: M
authors.lastname: Debatis
authors.forename: Shixia
authors.initials: S
authors.lastname: Wang
authors.forename: Shan
authors.initials: S
authors.lastname: Lu
authors.forename: Timothy F
authors.initials: TF
authors.lastname: Kowalik
mesh_terms: D002460:Cell Line; D003587:Cytomegalovirus / Q000235:genetics* / Q000502:physiology; D003586:Cytomegalovirus Infections / Q000235:genetics / Q000378:metabolism / Q000821:virology*; D004249:DNA Damage; D004259:DNA-Directed DNA Polymerase / Q000235:genetics* / Q000378:metabolism; D015536:Down-Regulation; D015967:Gene Expression Regulation, Viral; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics* / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D000071156:Promyelocytic Leukemia Protein; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D014779:Virus Replication
chemical_list: C069186:IE2 protein, Cytomegalovirus; D017874:Immediate-Early Proteins; D009687:Nuclear Proteins; D000071156:Promyelocytic Leukemia Protein; D034741:RNA, Small Interfering; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C109367:UL54 protein, Human herpesvirus 5; D014764:Viral Proteins; C070576:PML protein, human; D004259:DNA-Directed DNA Polymerase