title: CRISPR/Cas9-Mediated Genome Editing of Herpesviruses Limits Productive and Latent Infections.
abstract: Herpesviruses infect the majority of the human population and can cause significant morbidity and mortality. Herpes simplex virus (HSV) type 1 causes cold sores and herpes simplex keratitis, whereas HSV-2 is responsible for genital herpes. Human cytomegalovirus (HCMV) is the most common viral cause of congenital defects and is responsible for serious disease in immuno-compromised individuals. Epstein-Barr virus (EBV) is associated with infectious mononucleosis and a broad range of malignancies, including Burkitt's lymphoma, nasopharyngeal carcinoma, Hodgkin's disease, and post-transplant lymphomas. Herpesviruses persist in their host for life by establishing a latent infection that is interrupted by periodic reactivation events during which replication occurs. Current antiviral drug treatments target the clinical manifestations of this productive stage, but they are ineffective at eliminating these viruses from the infected host. Here, we set out to combat both productive and latent herpesvirus infections by exploiting the CRISPR/Cas9 system to target viral genetic elements important for virus fitness. We show effective abrogation of HCMV and HSV-1 replication by targeting gRNAs to essential viral genes. Simultaneous targeting of HSV-1 with multiple gRNAs completely abolished the production of infectious particles from human cells. Using the same approach, EBV can be almost completely cleared from latently infected EBV-transformed human tumor cells. Our studies indicate that the CRISPR/Cas9 system can be effectively targeted to herpesvirus genomes as a potent prophylactic and therapeutic anti-viral strategy that may be used to impair viral replication and clear latent virus infection.
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Ferdy R
authors.initials: FR
authors.lastname: van Diemen
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Elisabeth M
authors.initials: EM
authors.lastname: Kruse
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Marjolein J G
authors.initials: MJ
authors.lastname: Hooykaas
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Carlijn E
authors.initials: CE
authors.lastname: Bruggeling
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Anita C
authors.initials: AC
authors.lastname: Schürch
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Petra M
authors.initials: PM
authors.lastname: van Ham
authors.affiliation: Department of Ophthalmology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Saskia M
authors.initials: SM
authors.lastname: Imhof
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Monique
authors.initials: M
authors.lastname: Nijhuis
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Emmanuel J H J
authors.initials: EJ
authors.lastname: Wiertz
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: Robert Jan
authors.initials: RJ
authors.lastname: Lebbink
mesh_terms: D064113:CRISPR-Cas Systems / Q000235:genetics*; D002460:Cell Line; D003587:Cytomegalovirus / Q000235:genetics*; D000072669:Gene Editing / Q000379:methods*; D016679:Genome, Viral; D006564:Herpesviridae / Q000235:genetics*; D006566:Herpesviridae Infections / Q000235:genetics*; D018259:Herpesvirus 1, Human; D006801:Humans; D016133:Polymerase Chain Reaction; D017735:Virus Latency / Q000235:genetics