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closeLack of lytic antigen does not indicate latency
Posted by virusdoc on 01 Jul 2007 at 19:58 GMT
These were presumably latently infected
http://plosone.org/article/info:doi/10.1371/journal.pone.0000560#article1.body1.sec2.sec5.p3
The authors presume that macrophages displaying some evidence of infection (expression of GFP from the viral genome) but not full lytic infection (lack of orf65 staining) are latently infected. This is not the only possibility. Other possibilities include abortive infection or infection in the face of an antiviral response. The latter is most probable, since Gresser's group demonstrated over 20 years ago that peritoneal macrophages (the cells used in this assay) are in an interferon-induced antiviral state for several days after explant (J. Virol 57:456; 1986). It is likely that the authors of this manuscript are simply studying cells which are antiviral due to in vivo interferon exposure, and thus non-permissive for infection. This state is not equivalent to latency, and should not be confused with it.
Reference:
http://www.ncbi.nlm.nih.gov/sites/entrez?Db=pubmed&Cmd=ShowDetailView&TermToSearch=3003378
RE: Lack of lytic antigen does not indicate latency
pgs27 replied to virusdoc on 10 Jul 2007 at 10:40 GMT
We discounted this possibility for the following reasons.
1. We get exactly the same results with ex vivo peritoneal macrophages and with RAW264.7 cells, a cloned macrophage line that is propagated in vitro. In vivo effects therefore do not explain the results.
2. Peritoneal macrophages do not resist lytic infection. Over a few days, they all become nuclear capsid positive. This is inconsistent with abortive infection, etc.
3. In our hands, interferon pre-treatment of cells blocks both lytic gene expression and HCMV IE1-driven eGFP from the MHV-68 genome. This is different to the non-lytic eGFP expression we observe in macrophages. Interferon-gamma treatment of macrophages after infection inhibits lytic infection but enhances HCMV IE1-driven eGFP expression, much as we reported for LPS. It would therefore seem that eGFP+ expression and lytic infection are different states.
It is difficult to study the non-lytic state with wild-type MHV-68 because it is not stably maintained. But the obvious interpretation of infection followed by a non-lytic state followed by lytic infection would be latency.
4. ORF50-deficient MHV-68 cannot go lytic. But it still expresses eGFP from the HCMV IE1 promoter in RAW264.7 cells.
RE: Lack of lytic antigen does not indicate latency
rosagt replied to virusdoc on 11 Jul 2007 at 13:20 GMT
In addition to pgs' reasons, we discounted abortive infection or infection in face of an antiviral state because a latency deficient MHV-68, the M50 (May JS, J Gen Virol. 2004 Jan;85(Pt 1):137-46.), easily infects macrophages (both peritoneal macrophages and RAW 264.7).
Infection with the M50 virus yields a much higher percentage of nucleocapsid+ cells because it is more lytic and it also expresses eGFP from the HCMV IE1 promoter. With the M50, all eGFP+ cells are nucleocapsid+. If anything, eGFP expression is lower, presumably due to the competition for protein synthesis. Thus, eGFP expression was not associated with abortive infection.
Additionally, with the same MOI, the percentage of eGFP+ cells infected with wild-type, treated with LPS (as shown in the paper), is similar to the percentage of nucleocapsid+ cells infected with the M50.