Day 72: What’s on the virus and what’s needed to make the virus
January 21, 2008
I’ve just spent the last hour comparing what’s found in HIV virions with what host factors the virus needs to produce virions. Chertova et al. published a great paper a few years ago in the Journal of Virology where they used mass spectrometry to determine what proteins are found in HIV particles produced in macrophages (primary cells). Brass and colleagues recently published a study in Science in which they used an RNAi knockdown screen to determine what host proteins are needed for viral production in HeLa and TZM-Bl cells (cell lines). It is very important to note that HeLas and TZM-Bls (which I believe are derived from HeLas, if memory serves) are very different cells from primary macrophages, so you might expect that HIV might have different requirements for each cell type and that some of the proteins found in one cell type may not even be present in the other types. Just to give you an idea of how different they are, macrophages are monocytic cells that are isolated from donors and can remain in culture for maybe a month (longer maybe?) and HeLa cells are epithelial cells originally derived from a cervical cancer tumor in 1951 at Hopkins. They’ve been around for over 50 years and they grow like weeds. That’s my caveat.
What I noticed between the two studies is that there is virtually no overlap in the proteins found. Now, some of this is because of the assays used—the Brass study failed to find some of the proteins that we know are needed for HIV production, so we know that the screen didn’t identify all the proteins involved in HIV production. Also, Brass ruled out any knockdowns that severely limited cell viability or growth; some of those proteins might be found in the virus. I won’t rule out the human factor either, since the Brass article uses protein abbreviations and the Chertova article uses full names (my eyes are only so good, folks). Still, the amazing lack of concordance is really striking. Of all the proteins found in each of the studies (~200 each), I could only find one host protein that is both necessary for viral production and is also found in the viral particle: KIFC3 kinesin. I don’t know anything about this protein specifically, except that it is a motor and probably is involved in viral protein trafficking. It’s also interesting that the mass spec paper identified a bunch of Rab and VPS proteins that are incorporated in the virions and the RNAi paper found members of these classes that are needed for production—just not the same proteins. This could be due to trafficking differences between the cell types, but I found it odd that none of the proteins from these classes overlapped in the two studies.
I am still not sure what this means. I have always felt that the host proteins found in the virion are probably important or necessary for viral production. However, after studying this in the lab for four years and having little-to-no success, maybe I need to reevaluate this stance. Certainly, the lack of correlation between these studies can be due to a number of factors. Also, the fact that host proteins in the virion may not be necessary for viral production may not mean they are not important in HIV disease. I would really like to see someone use these two assays in one paper to try and make some of these correlations. Even better, I wish someone would do an RNAi screen and produce virus, testing whether virus produced is less infectious. This might be a great way to produce a therapeutic vaccine.
M. Linde
January 26, 2008 at 2:43 pm
I can’t remember how the RNAi study was done (and am either too busy, or too lazy [pick one!]) to check — did they evaluate more than one round of viral replication? If not, then there’s not necessarily a contradiction; one might expect that if the virion particle brings in a host protein then it may not need that protein in its first round of replication, because it’s providing it.
What you’d predict in that case is that knocking down proteins incorporated in the virion would not affect replication at first — but that virions exiting from the knockdown cell would not efficiently infect another knockdown cell (but perhaps could infect a wild-type cell).
January 26, 2008 at 2:44 pm
… And looking again at your post I think you are making the same point about the “less infectious” virus. Sorry about that.