From: Josh Bembenek (jbembe@hotmail.com)
Date: Tue Apr 01 2003 - 16:21:03 EST
Preston-
"What seems to be going on here, at least as interpreted by the researchers,
is not site specific insertion, but selection for cells in which the virus
inserted near a particular gene. I'm not a gene therapy expert, but I think
it works like this. They isolate some pre-lymphocyte cells of a particular
type from the patient and transform them in mass with the engineered
retrovirus vector. They may then select for cells that have the vector
incorporated, based on some selectable marker in the vector. I'm not sure
that they would do that or not. They would do it if the transformation is
inefficient. But they didn't apparently clone from a single transformant
cell, since they assume in their analysis that the cells they injected back
into the patient had the vector inserted at various sites. After the cells
are in the patient, some of them behave well and go to the marrow and start
producing the desired corrected lymphocytes. However, it turns out that some
element(s) of this virus can interact "in cis" (if the virus inserts next to
or within) with this LMO1 gene to give a cell that is unregulated for growth
and/or programmed cell death. That rare cell grows out to give the leukemia.
The reason it happened in two patients is that they are transforming and
injecting enough cells to make it fairly likely that in each patient at
least one cell will have the insertion somewhere near the LMO1 gene. Of
course, it would be interesting to know if the two insertions occurred at
exactly the same site or only in the same vicinity, which is much more
likely. There are such things as hotspots for virus insertion, probably
based on chromatin structure or sites related to normal recombination. It's
possible something like that contributed here, but I believe that in general
retroviruses are not very site specific."
-This is exactly why I've been pondering this piece of evidence for a while.
Since viruses supposedly jump into the genome completely at random, the
only way to get viruses integrated into the same site in the genome is to
have an ancestor obtain the viral incorporation, then pass it on to the
progeny. This requires incorporation into the genome of Gametes, but within
one infected individual who knows the number of possible viral incorporation
into gametes, and then there's the issue of that gamete being passed on...
"As to your larger question on endogenous retroviruses at the same site in
different species, a mechanism corresponding to that above would require
that there be a selective advantage of the insertion at the same exact
location for both species."
-Why is this so? Two requirements for similar site insertions across
species come to mind: gametes must be infected, and similar variables must
lead to similar insertions (either virus has some unknown specificity for
something like non-methylated DNA or degree of chromatin
condensation/silencing...) If the insertions are neutral, they will be
retained in the genome, because they will simply piggyback with the
chromosome when DNA duplication occurs. I don't know of any virus element
editing mechanisms during genome duplication that would get rid of any
inserted virus. Also, are known viral insertions demonstrated to be
conferring selective advantage to the host?
There may be a few sites where there was a selective advantage, but that
would be dubious for the vast majority. Even in that case, you wouldn't
expect the insertion to occur at exactly the same site.
-Exactly why I find this 2/10 case for leukemia like symptoms in this gene
therapy trial case interesting. Also, this has only taken 3 years to
develop, whatever events trigger these symptoms may occur later in more of
the treated patients. This would appear to argue that the virus insertion
has a site preference.
Insertion at exactly the same site in humans and other primates is I think
known to be the case for some retrovirus insertions where the region has
been sequenced in more than one species. Other transposable elements,
particularly L1 elements and Alu elements are present in mammalian genomes
in much larger numbers than retroviruses (L1 > 10^5; Alu > 10^6) and thus
provide thousands more examples of the same thing. These elements are so
frequent in the genome that almost any segment over a few kb that is
sequenced in both human and chimp or gorilla will have examples of this.
Some particular insertions can be shown to be present only after a certain
point in primate evolution. That is, they may be present only in chimps,
gorilla and human and not in more primitive primates or other mammals.
-Elaborating on the question I am raising, this may indicate the range of
hosts for a particular virus infection instead of an evolutionary
relationship. The question is, are there any other ways to interpret the
data in general. I am trying to think of other considerations.
Consider the following from Sverdlov, "Retroviruses and Primate Evolution"
Bioessays 22:161-171:
"..since retroviruses can cause cross-species infections, and there are data
suggesting multiple ancient cross-species transmissions of retroviruses in
primates."
Referenced articles: j. virol 1995 69:7877-7887, virology 1997 238 212-220.
Additionally, since viruses mutate much more rapidly when not incorporated
into a host, it is possible that events of viral incoporation in gametes
occur within the same wave of "retroviral plague" if you will, and that the
resulting incorporations would look different due to the differences in
rapidly mutating viral strains being incorporated, not their divergence due
to evolution after integration (at the same time alot of the mutations
observed seem to compromise viral function....)
Josh
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