Susceptibility of (XMRV) to retroviral restriction factors

[RedRuth]

I thought this was worth its own thread as it has important implications for the type of cells XMRV can infect and replicate in.
Apparently both Human and Mouse tetherin/CD317 and APOBEC restrict XMRV efficiently and there doesn't seem to be any viral accessory proteins that counteract these factors (like Vpu - an HIV-1 accessory protein). This is important as PBMCs and all hematopoietic cells express tetherin/CD317. If XMRV is an infective Human virus it's not likely to replicate in these cells. I also know that alot of prostate cancer cell lines don't express tetherin.

Susceptibility of xenotropic murine leukemia virus-related virus (XMRV) to retroviral restriction factors
http://www.pnas.org/content/107/11/5166.full

Xenotropic murine leukemia virus-related virus (XMRV) is a recently discovered gammaretrovirus that has been linked to prostate cancer and chronic fatigue syndrome. This virus is therefore an important potential human pathogen and, as such, it is essential to understand its host cell tropism. Intriguingly, infectious virus has been recovered from patient-derived peripheral blood mononuclear cells. These cells express several antiviral restriction factors that are capable of inhibiting the replication of a wide range of retroviruses, including other gamma retroviruses. This raises the possibility that, similar to HIV, XMRV may have acquired resistance to restriction. We therefore investigated the susceptibility of XMRV to a panel of different restriction factors. We found that both human APOBEC3 and tetherin proteins are able to block XMRV replication. Expression of human TRIM5?, however, had no effect on viral infectivity. There was no evidence that XMRV expressed countermeasures to overcome restriction. In addition, the virus was inhibited by factors from nonhuman species, including mouse Apobec3, tetherin, and Fv1 proteins. These results have important implications for predicting the natural target cells for XMRV replication, for relating infection to viral pathogenicity and pathology, and for the design of model systems with which to study XMRV-related diseases.

 

[eric_s]

This paper is from 2010, are you sure there is not already a thread about it? I have read about it before.

 

[currer]

Thanks, Redruth,
This paper comes from last year, but I have a better chance of understanding it now. It will be interesting bedtime reading.
There may be something more recently published on this subject though, can you find anything?

 

[RedRuth]

I had a quick look and didn't see a thread then I did a better search and found it. The main reason I posted it though is because I have a research interest in some of the proteins but from a different perspective. But I can ask a mod to delete it, I hadn't seen the paper so I was interested in it.

 

[RedRuth]

you're welcome, there's usually a 'cited by' list so I'll have a look.

 

[eric_s]

I don't mind, but maybe it's better to continue the discussion in the already existing thread.

 

[RedRuth]

This is a very recent paper that also looks at restriction factors http://jvi.asm.org/cgi/content/full/85/10/4888 They infected activated PBMCs and cell lines that are +/- for APOBEC proteins with XMRV from 22Rv1 cells and looked at virus replication. A couple of points from the discussion

Our observation that XMRV replication and spread is severely restricted in PHA-activated PBMCs is a novel finding that has not so far been reported. These results are also inconsistent with the report by Lombardi et al. (28), which showed that after activation with PHA and IL-2, nearly all of the PBMCs from XMRV-positive patients become reactive to a monoclonal antibody against the MLV p30 Gag protein; these studies suggested that XMRV replicated and spread efficiently in CFS patients and was present in most of the activated PBMCs (28). In future studies, it will be important to compare the replication and spread of XMRV in PBMCs isolated from normal control donors and CFS patients.

and

Despite the potent restriction, the PBMCs could be infected with XMRV, as recently shown (17), and the infected PBMCs could produce virus particles; this is consistent with the mechanism of A3G/A3F-mediated inhibition of viral replication, which does not block virus entry or production but greatly reduces the infectivity of the virions (47, 61). We also observed G-to-A hypermutation of XMRV proviruses in PBMCs, indicating that virus produced from the PBMCs could reinfect the cells and generate proviruses; since there was no detectable increase in virus production, most of these reinfection events probably resulted in abortive infection or the generation of highly mutated proviral genomes that were unable to produce viral particles. The extent of hypermutation was variable between different donors and very low for donor 1, suggesting that replication and spread of XMRV in PBMCs from some donors may be very inefficient. These results indicate that hypermutation of XMRV proviral genomes at GG nucleotides, which indicate A3G substrate specificity, may serve as a useful marker for infection and replication of XMRV in human PBMCs.

Has the Mikovits lab ever reported hypermutated XMRV at GG postions?

 

[currer]

Hi Redruth,

The WPI are having trouble getting their research published so no-one knows.

Thanks for finding this.

I cant get access to read the whole paper, unfortunately.

 

[alex3619]

Hi RedRuth, Judy Mikovits did discuss hypermutation at the New York Academy of Sciences workshop some months ago, iirc. They find it all the time, but it does not seem to stop the virus so much as slow it. I do not know if the data was published though. Bye, Alex

 

[Mark]

Regrettably my recollection is hazy once again...but this does seem to be an important area.

My recollection of the earlier threads from a year or so ago is that there were apparently conflicting results, and that it rather looked to me as though APOBEC inhibits XMRV but also XMRV 'inhibits' APOBEC...I probably have this wrong but my take-home from that discussion was the suggestion that XMRV and the inhibition factors seemed to be locked in battle...

The hypermutation processes also seemed very significant, and the suggestion that XMRV may be resistant to APOBEC's mutation seemed most intriguing. My layman's understanding (please forgive any inaccurate terminology) was this: APOBEC causes mutations in foreign DNA (from invading retroviruses), by switching (eg) G to A so as to inactivate them....but it has been suggested (but perhaps not published) that these induced mutations in XMRV happen to result in the same proteins, such that XMRV can in certain circumstances be resistant to the hypermutation process. In other words, after the mutations have taken place, the mutated sequence still replicates to produce XMRV again.

It's probably not quite as I've described, and I'm not sure whether that information was ever published (though I'm pretty sure there was one paper suggesting inhibition of APOBEC itself by XMRV under some circumstances), but: what intrigued me was that if this was indeed the case, then this process would seem to be highly selective for the XMRV sequence, and I wondered whether this could explain the low sequence diversity of XMRV. Such a process would seem to be highly selective of the retroviral sequence, because only very specific sequences would survive the editing process.

I'd be very interested to hear from RedRuth whether this rough hypothesis makes any sense.

The other interesting angle was that there may be genetic deficiencies in the APOBEC genes which could explain why some people are more vulnerable to XMRV infection. Once again I'm hazy on the detail, but I seem to remember this APOBEC weakness linking up well with other ME/CFS research.

 

[*GG*]

Where is the other thread that this is being discussed on?

GG

 

[Jemal]

My recollection of the earlier threads from a year or so ago is that there were apparently conflicting results, and that it rather looked to me as though APOBEC inhibits XMRV but also XMRV 'inhibits' APOBEC...I probably have this wrong but my take-home from that discussion was the suggestion that XMRV and the inhibition factors seemed to be locked in battle...

This I remember reading as well. XMRV even spreading (or holding?) into cells that were supposedly protected? But I remember reading about it on other websites I think.

 

[Jemal]

Where is the other thread that this is being discussed on?

GG

This is that thread I think. From december last year:
http://forums.phoenixrising.me/show...C3-proteins-can-inhibit-XMRV&highlight=APOBEC

 

[RedRuth]

My recollection of the earlier threads from a year or so ago is that there were apparently conflicting results, and that it rather looked to me as though APOBEC inhibits XMRV but also XMRV 'inhibits' APOBEC...I probably have this wrong but my take-home from that discussion was the suggestion that XMRV and the inhibition factors seemed to be locked in battle.

You mean the Groom et al paper in the OP? APOBEC and tetherin both efficiently restrict XMRV and they found no evidence that XMRV expresses any counter measures (like the vpu protein of HIV) The inference being if XMRV is infecting PBMCs then the host restriction factors expressed by these cells (tetherin and APOBEC) would stop the virus replicating (or to be more precise releasing infective virus particles). Maybe you mean TRIM5 which had no effect on XMRV. This contradicts the Lombardi paper.

The hypermutation processes also seemed very significant, and the suggestion that XMRV may be resistant to APOBEC's mutation seemed most intriguing

The point of both papers is that XMRV isn't resistant to APOBEC or tetherin.

 

[RedRuth]

This I remember reading as well. XMRV even spreading (or holding?) into cells that were supposedly protected? But I remember reading about it on other websites I think.

The Groom et al paper shows that XMRV is restricted efficiently by both APOBEC and tetherin but not by TRIM5alpha. And as the say Taken together, these data demonstrate that wild-type XMRV does not encode a human, primate, or murine tetherin antagonist and is, thus, highly sensitive to these restriction factors.

This was all done in tissue culture cells so the more recent paper went on to look at activated PBMCs infected with XMRV isolated from 22Rv1 cells. Again they found that APOBEC efficiently restricted XMRV but that the hypermutation was variable, however PBMCs aslo express tetherin so there's another line of defence. This directly contradicts the Lomabardi paper as they point out. Our observation that XMRV replication and spread is severely restricted in PHA-activated PBMCs is a novel finding that has not so far been reported. These results are also inconsistent with the report by Lombardi et al.

They did though find that although there was efficient restriction of XMRV they could infect a co culture of canine cells meaning that there were some viable virus particles present. How physiologically relevant this is I couldn't say (I'm not a virologist) They specualte that PBMCs could be a reservoir or carrier rather than the site of replication.

EDIT: RE what you said about XMRV 'holding on to cells', this may be a misunderstanding of how tetherin works. Like it's name suggests it tethers nascent virus particles to the host cell membrane as they are trying to bud from the host cell (after replication). The viral particles are then re internalised and degraded by the host cell lysosomes. There's some great EM images of HIV particles 'tethered' to cells in the original Nature paper.

 

[RedRuth]

That's a different paper. The papers I've cited

Susceptibility of xenotropic murine leukemia virus-related virus (XMRV) to retroviral restriction factors http://www.pnas.org/content/107/11/5166.full

Severe Restriction of Xenotropic Murine Leukemia Virus-Related Virus Replication and Spread in Cultured Human Peripheral Blood Mononuclear Cells http://jvi.asm.org/cgi/content/full/85/10/4888

 

[currer]

Dr Mikovits remarks about restriction of XMRV are on the thread "XMRV & Blood Supply Webinar March 29" on the media page.
Alex put up a summary on page 5. The webinar is not publicly available now, so we cannot go back to it.

On another tack, the webinar mentioned "aerosolizing" suggesting that XMRV might be able to be airborne.
This is also discussed in this paper - the correspondence about the biosafety level 2 recombinant retrovirus. The Germans who authored the paper say that it is unclear how MLV contaminants spread. They suggest formation of aerosols during the splitting of a contaminated culture and subsequent contamination of cultures splitted in the same tissue culture hood via aerosols. http://www.retrovirology.com/content/6/1/86/comments

 

[Jemal]

Dr Mikovits remarks about restriction of XMRV are on the thread "XMRV & Blood Supply Webinar March 29" on the media page.
Alex put up a summary on page 5. The webinar is not publicly available now, so we cannot go back to it.

On another tack, the webinar mentioned "aerosolizing" suggesting that XMRV might be able to be airborne.
This is also discussed in this paper - the correspondence about the biosafety level 2 recombinant retrovirus. The Germans who authored the paper say that it is unclear how MLV contaminants spread. They suggest formation of aerosols during the splitting of a contaminated culture and subsequent contamination of cultures splitted in the same tissue culture hood via aerosols. http://www.retrovirology.com/content/6/1/86/comments

Why do I keep running into Dusty Miller?

 

[currer]

No but this perhaps suggests that XMRV could spread as an airbourne disease. The weakest link in the XMRV argument for me was always the epidemic outbreaks, as this suggested some other virus, an enterovirus perhaps.
But with the Zhang paper on the contamination problem in the lab, and this german correspondence, perhaps this retrovirus can spread through the air?
Which will be surprising.

 

[RedRuth]

Hi RedRuth, Judy Mikovits did discuss hypermutation at the New York Academy of Sciences workshop some months ago, iirc. They find it all the time, but it does not seem to stop the virus so much as slow it. I do not know if the data was published though. Bye, Alex

Did she mean APOBEC induced hypermutation? APOBEC is only one of the lines of defence against retroviral infection though. BTW, someone said you had posted a summary of what she had to say about restriction factors but I can't find it. Any chance you could point me in the right direction?