Clonal Immortalized Human Glial Cell Lines Support Varying Levels of JC Virus Infection due to Differences in Cellular Gene Expression

Abstract

JC virus (JCV) is a ubiquitous human polyomavirus that causes the demyelinating disease Progressive Multifocal Leukoencephalopathy (PML). JCV replicates in limited cell types in culture, predominantly in human glial cells. Following introduction of a replication defective SV40 mutant that expressed large T protein into a heterogeneous culture of human fetal brain cells, multiple phenotypes became immortalized (SVG cells). A subset of SVG cells could support JCV replication. In the current study, clonal cell lines were selected from the original SVG cell culture. The 5F4 clone showed low levels of viral growth. The 10B1 clone was highly permissive for JCV DNA replication and gene expression and supported persistent and stable JCV infection over months in culture. Microarray analysis revealed that viral infection did not significantly change gene expression in these cells. More resistant 5F4 cells expressed high levels of transcription factors known to inhibit JCV transcription. Interestingly, 5F4 cells expressed high levels of RNA of markers of radial glia and 10B1 cells had high expression of markers of immature glial cells and activation of transcription regulators important for stem/progenitor cell self-renewal. These SVG-derived clonal cell lines provide a biologically relevant model to investigate cell type differences in JCV host range and pathogenesis, as well as neural development. Several transcription regulators were identified which may be targets for therapeutic modulation of expression to abrogate JCV replication in PML patients. Additionally, these clonal cell lines can provide a consistent culture platform for testing therapies against JCV infection of the central nervous system.

The Human Fetal Glial Cell Line SVG p12 Contains Infectious BK Polyomavirus

“…This is supported by a recent study showing that SVG cells give rise to clonal cell lines with different phenotypes (34). We therefore speculate that subcloning of SVG p12 cells might give rise to uncontaminated cell lines.…”

“…The SVG p12 cell line is one of few human cell lines available for propagation of JCPyV (6,34). As such, a number of previous JCPyV studies, including drug efficacy studies (27,36) as well as production of antigen for antibody detection assays (26), have utilized SVG or subclones of these cells.…”

“…SVG p12, SVG, and subclones of SVG have been used in several JCPyV studies (26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41) and in more than 30 other studies in which cells of neural origin were required. Unfortunately, the source of the SVG cells was not clearly specified for some of these studies.…”

p53 isoforms regulate astrocyte-mediated neuroprotection and neurodegeneration

“…Treatment with bafilomycin A1 resulted in the upregulation of Δ133p53 in senescent astrocytes (Figure 2b), suggesting that autophagic degradation of Δ133p53 contributes to its downregulation in these cells. The stabilization of Δ133p53 by bafilomycin A1 was also observed in an immortalized human astrocyte cell line 39,40 (Supplementary Figure S2A). Knockdown of p62/SQSTM1, which is a ubiquitin-binding adaptor specifically functioning in the selective autophagy pathway, 41 also stabilized Δ133p53 (Figure 2c), further supporting the degradation of Δ133p53 via selective autophagy.…”

“…After 3 weeks, cells were transfected using calcium phosphate precipitation with a plasmid DNA containing the SV40 mutant with a deletion of its origin of replication (pMK16). 39,40 H358 cells were purchased from American Type Culture Collection (Manassas, VA, USA).…”