Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States; Harvard Stem Cell Institute, Harvard University, Cambridge, United States; Department of Obstetrics and Gynecology, Division of Reproductive Endocrinology and Infertility, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, United States
Pierce Eggan
Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States; Harvard Stem Cell Institute, Harvard University, Cambridge, United States
Magdalena Angelova
Department of Microbiology, Blavatnik Institute, Harvard Medical School, Boston, United States
Rory Kirchner
Department of Biostatistics, Harvard TH Chan School of Public Health, Boston, United States
Kevin C Eggan
Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States; Harvard Stem Cell Institute, Harvard University, Cambridge, United States; Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, United States
Herpes simplex virus (HSV) establishes lifelong latent infection and can cause serious human disease, but current antiviral therapies target lytic but not latent infection. We screened for sgRNAs that cleave HSV-1 DNA sequences efficiently in vitro and used these sgRNAs to observe the first editing of quiescent HSV-1 DNA. The sgRNAs targeted lytic replicating viral DNA genomes more efficiently than quiescent genomes, consistent with the open structure of lytic chromatin. Editing of latent genomes caused short indels while editing of replicating genomes produced indels, linear molecules, and large genomic sequence loss around the gRNA target site. The HSV ICP0 protein and viral DNA replication increased the loss of DNA sequences around the gRNA target site. We conclude that HSV, by promoting open chromatin needed for viral gene expression and by inhibiting the DNA damage response, makes the genome vulnerable to a novel form of editing by CRISPR-Cas9 during lytic replication.
