Shared ancestry of herpes simplex virus 1 strain Patton with recent clinical isolates from Asia and with strain KOS63

Abstract

Herpes simplex virus 1 (HSV-1) is a widespread pathogen that persists for life, replicating in surface tissues and establishing latency in peripheral ganglia. Increasingly, molecular studies of latency use cultured neuron models developed using recombinant viruses such as HSV-1 GFP-US11, a derivative of strain Patton expressing green fluorescent protein (GFP) fused to the viral US11 protein. Visible fluorescence follows viral DNA replication, providing a real time indicator of productive infection and reactivation. Patton was isolated in Houston, Texas, prior to 1973, and distributed to many laboratories. Although used extensively, the genomic structure and phylogenetic relationship to other strains is poorly known. We report that wild type Patton and the GFP-US11 recombinant contain the full complement of HSV-1 genes and differ within the unique regions at only eight nucleotides, changing only two amino acids. Although isolated in North America, Patton is most closely related to Asian viruses, including KOS63.

Direct RNA sequencing on nanopore arrays redefines the transcriptional complexity of a viral pathogen

“…To evaluate the reproducibility of direct RNA sequencing using nanopore arrays, total RNA was prepared from two biological replicates of normal human dermal fibroblasts (NHDF) infected with HSV-1 GFP-Us11 strain Patton (hereafter HSV-1 Patton) 23,24 for 18 h. Sequencing libraries were generated from the poly(A) + RNA fraction and sequenced on a MinION MkIb with R9.4 flow cell with a run time of 18 h, yielding ~380,000 (replicate #1) and 218,000 (replicate #2) nanopore sequence reads (Fig. 1a, Supplementary Table 1), which were then aligned against the human transcriptome and HSV-1 strain 17 syn + annotated reference sequence using the splice-aware aligner MiniMap2 25 .…”

Native RNA sequencing on nanopore arrays redefines the transcriptional complexity of a viral pathogen

“…To evaluate the reproducibility of native RNA sequencing (nRNA-seq) using nanopore arrays we prepared total RNA from two biological replicates of normal human dermal fibroblasts (NHDF) infected with HSV-1 GFP-Us11 strain Patton (hereafter HSV-1 Patton) 22, 23 for 18 hours (Fig. 1a, Supplementary Fig.…”

Topoisomerase 2β-dependent nuclear DNA damage shapes extracellular growth factor responses through AKT phosphorylation dynamics to control virus latency

“…Sympathetic neurons from dissociated superior cervical ganglia (SCG) neurons from E21 rat embryos can be cultured as a pure population of cells that depend upon the trophic factor NGF (Glebova and Ginty, 2005). To determine whether genotoxic stress can affect HSV-1 latency in SCG neurons, we utilized a primary neuronal culture model system for establishing HSV-1 latency in vitro and employed a wildtype HSV-1 strain expressing the enhanced Green Fluorescent Protein (GFP) fused to the Us11 true-late (g2) protein (GFP-Us11) that can be detected during reactivation in living neurons (Benboudjema et al, 2003;Camarena et al, 2010;Kobayashi et al, 2012a;Linderman et al, 2017;Pourchet et al, 2017) (Figure 1A). Treatment of HSV-1 latently-infected neurons with either Bleomycin (DSB inducer, radiomimetic) or Etoposide (DSB inducer, TOP2 inhibitor) caused viral reactivation (productive viral replication leading to GFP-positive wells) at levels comparable to treatment with LY294002 (PI 3-kinase inhibitor, positive control).…”