by Jonathan Latham and Allison Wilson
January 21, 2013
How should a regulatory agency announce they have discovered
something potentially very important about the safety of products
they have been approving for over twenty years?
In the course of analysis to identify potential allergens in GMO
crops, the European Food Safety Authority (EFSA) has
belatedly discovered that the most common genetic regulatory
sequence in commercial GMOs also encodes a significant fragment of a
viral gene (Possible
Consequences of The Overlap Between The CaMV 35S Promoter Regions in
Plant Transformation Vectors Used and The Viral Gene VI in
Transgenic Plants - Podevin
and du Jardin 2012). This finding has serious ramifications for crop
biotechnology and its regulation, but possibly even greater ones for
consumers and farmers.
This is because there are clear
indications that this viral gene (called Gene VI) might not be safe
for human consumption. It also may disturb the normal functioning of
crops, including their natural pest resistance.
Cauliflower Mosaic Virus
What Podevin and du Jardin discovered is that of the 86 different
transgenic events (unique insertions of foreign DNA) commercialized
to-date in the United States 54 contain portions of Gene VI
They include any with a widely used gene
regulatory sequence called the CaMV 35S promoter (from the
cauliflower mosaic virus; CaMV). Among the affected transgenic
events are some of the most widely grown GMOs, including Roundup
Ready soybeans (40-3-2) and MON810 maize.
They include the controversial NK603
maize recently reported as causing tumors in rats (Seralini et al.
The researchers themselves concluded that the presence of segments
of Gene VI,
“might result in unintended
They reached this conclusion because
similar fragments of Gene VI have already been shown to be active on
their own (e.g. De Tapia et al. 1993). In other words, the EFSA
researchers were unable to rule out a hazard to public health or the
In general, viral genes expressed in plants raise both agronomic and
human health concerns (reviewed in Latham and Wilson 2008). This is
because many viral genes function to disable their host in order to
facilitate pathogen invasion. Often, this is achieved by
incapacitating specific anti-pathogen defenses.
Incorporating such genes could clearly
lead to undesirable and unexpected outcomes in agriculture.
Furthermore, viruses that infect plants are often not that different
from viruses that infect humans.
For example, sometimes the genes of
human and plant viruses are interchangeable, while on other
occasions inserting plant viral fragments as transgenes has caused
the genetically altered plant to become susceptible to an animal
virus (Dasgupta et al. 2001).
Thus, in various ways, inserting viral
genes accidentally into crop plants and the food supply confers a
significant potential for harm.
The original discovery by Podevin and du Jardin (at
EFSA) of Gene VI in commercial GMO crops must have presented
regulators with sharply divergent procedural alternatives.
recall all CaMV Gene
VI-containing crops (in Europe that would mean revoking
importation and planting approvals)
undertake a retrospective risk
assessment of the CaMV promoter and its Gene VI sequences
and hope to give it a clean bill of health
It is easy to see the attraction for
EFSA of option two.
Recall would be a massive political and
financial decision and would also be a huge embarrassment to the
regulators themselves. It would leave very few GMO crops on the
market and might even mean the end of crop biotechnology.
Regulators, in principle at least, also have a third option to gauge
the seriousness of any potential GMO hazard.
GMO monitoring, which is required by EU
regulations, ought to allow them to find out if deaths, illnesses,
or crop failures have been reported by farmers or health officials
and can be correlated with the Gene VI sequence.
Unfortunately, this particular avenue of
enquiry is a scientific dead end. Not one country has carried
through on promises to officially and scientifically monitor any
hazardous consequences of GMOs.(1)
Unsurprisingly, EFSA chose option two. However, their investigation
resulted only in the vague and unreassuring conclusion that Gene VI
“might result in unintended phenotypic changes” (Podevin and du
Jardin 2012). This means literally, that changes of an unknown
number, nature, or magnitude may (or may not) occur.
It falls well short of the solid
scientific reassurance of public safety needed to explain why EFSA
has not ordered a recall.
Can the presence of a fragment of virus DNA really be that
significant? Below is an independent analysis of Gene VI and its
known properties and their safety implications.
This analysis clearly illustrates the
Functions of Gene VI
Gene VI, like most plant viral genes, produces a protein that is
It has four (so far) known roles in the
viral infection cycle.
The first is to participate in
the assembly of virus particles. There is no current data to
suggest this function has any implications for biosafety.
The second known function is to
suppress anti-pathogen defenses by inhibiting a general
cellular system called RNA silencing (Haas et al. 2008).
Thirdly, Gene VI has the highly
unusual function of transactivating (described below) the
long RNA (the 35S RNA) produced by CaMV (Park et al. 2001).
Fourthly, unconnected to these
other mechanisms, Gene VI has very recently been shown to
make plants highly susceptible to a bacterial pathogen (Love
et al. 2012).
Gene VI does this by interfering with a
common anti-pathogen defense mechanism possessed by plants.
These latter three functions of Gene VI
(and their risk implications) are explained further below:
1) Gene VI Is an Inhibitor of RNA
RNA silencing is a mechanism
for the control of gene expression at the level of RNA abundance
It is also an important antiviral
defense mechanism in both plants and animals, and therefore most
viruses have evolved genes (like Gene VI) that disable it (Dunoyer
and Voinnet 2006).
Gene VI (upper
left) precedes the start of the 35S RNA
This attribute of Gene VI raises two obvious biosafety concerns:
1) Gene VI will lead to aberrant
gene expression in GMO crop plants, with unknown
2) Gene VI will interfere with
the ability of plants to defend themselves against viral
There are numerous experiments
showing that, in general, viral proteins that disable gene
silencing enhance infection by a wide spectrum of viruses
(Latham and Wilson 2008).
2) Gene VI Is a Unique
Transactivator of Gene Expression
Multicellular organisms make
proteins by a mechanism in which only one protein is produced by
each passage of a ribosome along a messenger RNA (mRNA).
Once that protein is completed the
ribosome dissociates from the mRNA. However, in a CaMV-infected
plant cell, or as a transgene, Gene VI intervenes in this
process and directs the ribosome to get back on an mRNA
(reinitiate) and produce the next protein in line on the mRNA,
if there is one.
This property of Gene VI enables
Cauliflower Mosaic Virus to produce multiple proteins from a
single long RNA (the 35S RNA). Importantly, this function of
Gene VI (which is called transactivation) is not limited to the
35S RNA. Gene VI seems able to transactivate any cellular mRNA (Futterer
and Hohn 1991; Ryabova et al. 2002).
There are likely to be thousands of
mRNA molecules having a short or long protein coding sequence
following the primary one. These secondary coding sequences
could be expressed in cells where Gene VI is expressed.
The result will presumably be
production of numerous random proteins within cells. The
biosafety implications of this are difficult to assess. These
proteins could be allergens, plant or human toxins, or they
could be harmless.
Moreover, the answer will differ for
each commercial crop species into which Gene VI has been
3) Gene VI Interferes with Host
A very recent finding, not
known by Podevin and du Jardin, is that Gene VI has a second
mechanism by which it interferes with plant anti-pathogen
defenses (Love et al. 2012).
It is too early to be sure about the
mechanistic details, but the result is to make plants carrying
Gene VI more susceptible to certain pathogens, and less
susceptible to others.
Obviously, this could impact
farmers, however the discovery of an entirely new function for
gene VI while EFSA’s paper was in press, also makes clear that a
full appraisal of all the likely effects of Gene VI is not
Is There a
Direct Human Toxicity Issue?
When Gene VI is intentionally expressed in transgenic plants, it
causes them to become chlorotic (yellow), to have growth
deformities, and to have reduced fertility in a dose-dependent
manner (Ziljstra et al 1996).
Plants expressing Gene VI also show gene
expression abnormalities. These results indicate that, not
unexpectedly given its known functions, the protein produced by Gene
VI is functioning as a toxin and is harmful to plants (Takahashi et
Since the known targets of Gene VI
activity (ribosomes and gene silencing) are also found in human
cells, a reasonable concern is that the protein produced by Gene VI
might be a human toxin.
This is a question that can only be
answered by future experiments.
Is Gene VI
Protein Produced in GMO Crops?
Given that expression of Gene VI is likely to cause harm, a crucial
issue is whether the actual inserted transgene sequences found in
commercial GMO crops will produce any functional protein from the
fragment of Gene VI present within the CaMV sequence.
There are two aspects to this question. One is the length of Gene VI
accidentally introduced by developers.
This appears to vary but most of the 54
approved transgenes contain the same 528 base pairs of the CaMV 35S
promoter sequence. This corresponds to approximately the final third
of Gene VI. Deleted fragments of Gene VI are active when expressed
in plant cells and functions of Gene VI are believed to reside in
this final third.
Therefore, there is clear potential for
unintended effects if this fragment is expressed (e.g. De Tapia et
al. 1993; Ryabova et al. 2002; Kobayashi and Hohn 2003).
The second aspect of this question is what quantity of Gene VI could
be produced in GMO crops?
Once again, this can ultimately only be
resolved by direct quantitative experiments. Nevertheless, we can
theorize that the amount of Gene VI produced will be specific to
each independent insertion event.
This is because significant Gene VI
expression probably would require specific sequences (such as the
presence of a gene promoter and an ATG [a protein start codon]) to
precede it and so is likely to be heavily dependent on variables
such as the details of the inserted transgenic DNA and where in the
plant genome the transgene inserted.
Commercial transgenic crop varieties can also contain superfluous
copies of the transgene, including those that are incomplete or
rearranged (Wilson et al 2006).
These could be important additional
sources of Gene VI protein. The decision of regulators to allow such
multiple and complex insertion events was always highly
questionable, but the realization that the CaMV 35S promoter
contains Gene VI sequences provides yet another reason to believe
that complex insertion events increase the likelihood of a biosafety
Even direct quantitative measurements of Gene VI protein in
individual crop authorizations would not fully resolve the
scientific questions, however.
No-one knows, for example, what
quantity, location or timing of protein production would be of
significance for risk assessment, and so answers necessary to
perform science-based risk assessment are unlikely to emerge soon.
It is perhaps the most basic assumption in all of risk assessment
that the developer of a new product provides regulators with
accurate information about what is being assessed.
Perhaps the next most basic assumption
is that regulators independently verify this information. We now
know, however, that for over twenty years neither of those simple
expectations have been met. Major public universities, biotech
multinationals, and government regulators everywhere, seemingly did
not appreciate the relatively simple possibility that the DNA
constructs they were responsible for encoded a viral gene.
This lapse occurred despite the fact that Gene VI was not truly
hidden; the relevant information on the existence of Gene VI has
been freely available in the scientific literature since well before
the first biotech approval (Franck et al 1980).
We ourselves have offered specific
warnings that viral sequences could contain unsuspected genes
(Latham and Wilson 2008). The inability of risk assessment processes
to incorporate longstanding and repeated scientific findings is
every bit as worrisome as the failure to intellectually anticipate
the possibility of overlapping genes when manipulating viral
This sense of a generic failure is reinforced by the fact that this
is not an isolated event.
There exist other examples of
commercially approved viral sequences having overlapping genes that
were never subjected to risk assessment. These include numerous
commercial GMOs containing promoter regions of the closely related
virus figwort mosaic virus (FMV) which were not considered by
Podevin and du Jardin.
Inspection of commercial sequence data
shows that the commonly used FMV promoter overlaps its own Gene VI (Richins
et al 1987).
A third example is the virus-resistant
potato NewLeaf Plus (RBMT-22-82).
This transgene contains approximately 90% of the P0 gene of potato
leaf roll virus. The known function of this gene, whose existence
was discovered only after US approval, is to inhibit the
anti-pathogen defenses of its host (Pfeffer et al 2002).
Fortunately, this potato variety was
never actively marketed.
A further key point relates to the biotech industry and their
campaign to secure public approval and a permissive regulatory
This has led them to repeatedly claim,
firstly, that GMO technology is
precise and predictable
secondly, that their own
competence and self-interest would prevent them from ever
bringing potentially harmful products to the market
thirdly, to assert that only
well studied and fully understood transgenes are
It is hard to imagine a finding more
damaging to these claims than the revelations surrounding Gene VI.
Biotechnology, it is often forgotten, is not just a technology. It
is an experiment in the proposition that human institutions can
perform adequate risk assessments on novel living organisms. Rather
than treat that question as primarily a daunting scientific one, we
should for now consider that the primary obstacle will be overcoming
the much more mundane trap of human complacency and incompetence.
We are not there yet, and therefore this
incident will serve to reinforce the demands for GMO labeling in
places where it is absent.
Regulators Should Do Now
This summary of the scientific risk issues shows that a segment of a
poorly characterized viral gene never subjected to any risk
assessment (until now) was allowed onto the market.
This gene is currently present in
commercial crops and growing on a large scale. It is also widespread
in the food supply.
Even now that EFSA’s own researchers have belatedly considered the
risk issues, no one can say whether the public has been harmed,
though harm appears a clear scientific possibility. Considered from
the perspective of professional and scientific risk assessment, this
situation represents a complete and catastrophic system failure.
But the saga of Gene VI is not yet over.
There is no certainty that further
scientific analysis will resolve the remaining uncertainties, or
provide reassurance. Future research may in fact increase the level
of concern or uncertainty, and this is a possibility that regulators
should weigh heavily in their deliberations.
To return to the original choices before EFSA, these were either to
recall all CaMV 35S promoter-containing GMOs, or to perform a
retrospective risk assessment. This retrospective risk assessment
has now been carried out and the data clearly indicate a potential
for significant harm.
The only course of action consistent
with protecting the public and respecting the science is for EFSA,
and other jurisdictions, to order a total recall.
This recall should also include GMOs
containing the FMV promoter and its own overlapping Gene VI.
1) EFSA regulators might now be
regretting their failure to implement meaningful GMO monitoring.
It would be a good question for European politicians to ask EFSA
and for the board of EFSA to ask the GMO panel, whose job it is
to implement monitoring.
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Safety Group Blows Lid on...
‘Secret Virus’ Hidden in GMO Crops
by Anthony Gucciardi
February 7, 2013
Yet another disturbing reason has
emerged as to why you should be avoiding health-devastating
genetically modified organisms, and it may be one of the most
We know that GMO consumption has been
linked to a host of serious conditions, but one thing we are not so
sure about is the recent discovery of a
hidden viral gene deep within
genetically modified crops.
For years, GMOs have been consumed
knowingly and unknowingly around the globe, with Monsanto and the
United States government claiming that the altered franken crops are
perfectly safe despite very limited (or virtually none in some
cases) initial testing and scientists speaking out against the
One such danger that has actually not
been spoken about has been revealed in a recent report by a safety
watchdog group known as the European Food Safety Authority (EFSA).
Secret of Monsanto
In the EFSA report, which can be
read online, you can find (within the scientific wording) that
researchers discovered a previously unknown
viral gene that is known as ‘Gene
What’s concerning is that not only is
the rogue gene found in the most prominent GMO crops and about 63%
of GMO traits approved for use (54 out of 86 to be precise), but it
can actually disrupt the very biological functions within
Popular GMO crops such as
...were found to contain the
gene that induces physical mutations. NK603 maize, of
course, was also
linked to the development of
mass tumors in rats.
Independent Science News, Gene
VI also inhibits RNA silencing.
As you may know, RNA silencing has been pinpointed as vital for the
proper functioning of gene expression when it comes to RNA. Perhaps
more topically, it is a defense mechanism against viruses in plants
and animals alike. On the contrary, many viruses have developed
genes that disable this protective process. Independent Science News
reports that the Gene VI is one such gene.
Overall, there is a degree of knowledge
on Gene VI.
What we do know going by information within the report
is that the gene:
Helps to assemble virus
Inhibits the natural defense of
the cellular system
Produces proteins that are
Makes plants susceptible to
All of which are very significant
effects that should be studied in depth by an independent team of
scientists after GMO products are taken off the market
pending further research on the entire array of associated diseases.
And that does not even include the
effects we are unaware of.
This is yet another incident in which
Monsanto and other biotech companies are getting away with an
offense against the citizens of the world with (most likely) no
action taken by the United States government.
What we have seen, however, is nations
...taking a stand against Monsanto in
direct opposition to their disregard for public safety.
Russia, in fact, banned Monsanto’s GMO
after it was linked to mass tumors in rats.
As more and more dirty secrets come out
from the GMO industry at large, it gives further reason and more
support to remove GMOs as a whole from the food supply.