Inefficient establishment of KSHV latency suggests an additional role for continued lytic replication in Kaposi sarcoma pathogenesis
Adam Grundhoff, Don Ganem
Adam Grundhoff, Don Ganem
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Article Infectious disease

Inefficient establishment of KSHV latency suggests an additional role for continued lytic replication in Kaposi sarcoma pathogenesis

Abstract

Kaposi sarcoma–associated (KS-associated) herpesvirus (KSHV) infection is linked to the development of both KS and several lymphoproliferative diseases. In all cases, the resulting tumor cells predominantly display latent viral infection. KS tumorigenesis requires ongoing lytic viral replication as well, however, for reasons that are unclear but have been suggested to involve the production of angiogenic or mitogenic factors by lytically infected cells. Here we demonstrate that proliferating cells infected with KSHV in vitro display a marked propensity to segregate latent viral genomes, with only a variable but small subpopulation being capable of stable episome maintenance. Stable maintenance is not due to the enhanced production of viral or host trans-acting factors, but is associated with cis-acting, epigenetic changes in the viral chromosome. These results indicate that acquisition of stable KSHV latency is a multistep process that proceeds with varying degrees of efficiency in different cell types. They also suggest an additional role for lytic replication in sustaining KS tumorigenesis: namely, the recruitment of new cells to latency to replace those that have segregated the viral episome.

Authors

Adam Grundhoff, Don Ganem

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Absence of stable episomal DNA from long-term cultures of cells transfec...
Absence of stable episomal DNA from long-term cultures of cells transfected with TR reporter plasmids. (a) Aliquots of FACS-sorted BJAB cells transfected with the indicated reporter constructs were harvested at various time points after FACS sorting (indicated in days above the lanes) and subjected to Gardella gel analysis. Samples were derived from the same cultures shown in Figure 1e. As a control, untransfected BJAB cells were loaded in the lanes labeled BJAB. Plasmid DNA was detected with a probe specific for GFP. The band marked with an asterisk results from unspecific background hybridization, since it is also present in untransfected BJAB cells. (b) Episomal DNA was isolated from the same BJAB cultures described above by Hirt extraction and subjected to PCR amplification of the GFP cassette as described in Methods. (c) Gardella gel analysis of transfected BJAB cells cultures grown under antibiotic selection. BJAB cells were transfected with pGFP, pGTR4, or pGTR4:73 and cultured in the presence of G418. Aliquots of cells propagated for the time periods indicated above the lanes (in days) were examined for the presence of episomal DNA by Gardella analysis. Episomes were detected with a probe specific for the GFP cassette.