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Old 09-10-2003, 09:22 PM
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Default A Danger to the World's Food: Genetic Engineering and the Economic Interests of the Life Science

GE 101-35S (PA)

The CaMV Promoter Story
The Cauliflower Mosaic Viral Promoter - A Recipe for Disaster?
by Dr. Mae-Won Ho, author of the book Biotechnology: Dream
or Nightmare?

The story of CaMV promoter encapsulates and draws attention to the
hazardous nature of the genetic engineering process itself as well as the
foreign gene constructs created and released into the environment.
Prof. Joe Cummins of the University of Western Ontario was the first
scientist to question the safety of the cauliflower mosaic viral (CaMV)
promoter, which is in practically all GM crops currently grown commercially
or undergoing field trials. His initial concern was that the promoter could
recombine with other viruses to generate new disease-causing viruses. In
our paper, we review some recent findings which give further grounds for
concern, and have recommended the immediate withdrawal of all crops
and products containing the CaMV promoter.
Ref.: Ho, M.W., Ryan, A. and Cummins, J. (1999). The cauliflower mosaic
viral promoter - a recipe for disaster? Microbial Ecology in Health and
Disease (in press).

To begin with, a 'promoter' is a stretch of genetic material that acts as a
switch for turning genes on. Every gene needs a promoter in order to work,
or to become expressed. But the promoter is not a simple switch like that
for an electric light, for example, which has only two positions, either
fully on or fully off. Instead, the promoter has many different modules that
act as sensors and to enable it to respond, in ways we do not yet fully
understand, to different signals from other genes and from the environment,
which tell it when and where to switch on, by how much and for how long.
And under certain circumstances, the promoter may be silenced, so that it
is off all the time.

All in all, the role of the promoter of a normal gene in an organism is to
enable the gene to work appropriately in the extremely complex regulatory
circuits of the organism as a whole. The promoter associated with each of
the organism's own genes is adapted to its gene while the totality of all
the genes of the organism have been adapted to stay and work together
for millions, if not hundreds of millions of years. The genome of each
organism is organised in a certain way which is more or less constant
across the species so individuals within a species can freely interbreed.
Each species protects its integrity and remains genetically stable because
there are biological barriers that prevent distant species from interbreeding
or otherwise exchanging genetic material. Foreign DNA are generally
broken down or inactivated.

Genetic engineering attempts to break down these biological barriers
so genes can be arbitrarily transferred between species that would
never interbreed in nature. In order to do so, special tricks are needed.
When genetic engineers transfer foreign genes into an organism to make
a GMO, they also have to put a promoter in front of each of the genes
transferred, otherwise it would not work. The promoter plus the gene it
switches on constitutes a 'gene-expression cassette'. Many of the genes
are from bacteria and viruses, and the most commonly used promoter is
from the caulifower mosaic virus. Several gene-expression cassettes are
usually stacked, or linked in series, one or more of them will be genes
that code for antibiotic resistance, which will enable those cells that have
taken up the foreign genes to be selected with antibiotics. The stacked
cassettes are then spliced in turn into an artificial gene carrier or 'vector'.
The vector is generally made by joining together parts of viruses and other
infectious genetic parasites (plasmids and transposons) that cause diseases
or spread antibiotic and drug resistance genes.

In the case of plants, the most widely used vector is the 'T-DNA'
which is part of the tumour-inducing plasmid ('Ti plasmid') of
Agrobacterium, a soil bacterium that infects plants and give rise to
plant tumours or galls. The role of the vector is to smuggle genes into
cells that would otherwise exclude them. And more importantly, the
vector can jump into the cell's genome and so enable the gene-expression
cassettes it carries to become incorporated into the genetic material of the
cell. The genetic engineer cannot control where and in what form the vector
jumps into the genetic material of the cell, however. And this is where the
first unpredictable effects can arise. Each transgenic line is unique, and
gives rise to different unintended effects, and in the case of food, can
include unexpected toxins and allergens.

The foreign genetic material transferred to make a transgenic organism -
referred to as the 'transgenic DNA' or the 'construct' - is quite complicated.
It consists of new genes and new combinations of genes - from diverse
species and their genetic parasites - which have never existed in nature.
Such chimaeric constructs are already known to be structurally unstable,
that is, they are prone to make and break and rearrange. It is to be
expected that such structural instability can only increase when the
construct is introduced, by a totally hit or miss process, into a new
genome. Transgenic instability is a well-known problem for the industry.
Transgenic lines often do not breed true (see Srivastava et al, 1999, in
item #3 below).

Why use a promoter from a virus such as the CaMV? A virus is a genetic
parasite that has the capability to infect the cell and hi-jack the cell to
make many copies of itself in a short period of time. Its promoter is
therefore very aggressive and hence popular with genetic engineers, as it
effectively makes the gene placed next to it turn on full blast, at perhaps
a thousand times the volume of any of the organism's own gene. Having it in
the genome is rather like having the loudest phrase of a heavy-metal piece
played with the most powerful amplifier simultaneously over and over again
throughout a live performance of a Mozart concerto. What the CaMV
promoter actually does is to place the foreign gene outside the normal
regulatory circuits of the host organism, subjecting the host organism
effectively to a permanent metabolic stress. This will multiply the unintended,
unpredictable effects, which are legion in transgenic organisms. It may also
be another reason why transgenic lines are notoriously unstable (Finnegan, J.
& McElroy, D. 1994, Bio/Technology 12, 883). The organism generally
reacts to the presence of foreign genetic material by breaking it down or
inactivating it. Even after the genetic material is incorporated into the genome,
it can silence the foreign genes so that they are no longer expressed (see
Item #3 below).

The key recent finding, which provoked our review, was the report (Kohli et
al, (1999) The Plant Journal 17, 591) that the CaMV promoter contains a
'recombination hotspot' - a site where the DNA tends to break and join up
with other DNA, thus changing the combination and arrangement of genes.
Around the hotspot are several short stretches or modules for binding
various enzymes, all of which are also involved in recombination , ie,
breaking and joining DNA. Furthermore, the CaMV promoter recombination
hotspot bears a strong resemblance to the borders of the T-DNA vector
carrying the transgenes, which are also known to be prone to recombination.
It is that which enables the vector to invade the cell's genome in the first

The aim of our original paper, restated explicitly in our official rebuttal,
was to review the relevant findings and, in particular, to point out the
implications, which the researchers themselves are unwilling or unable to
draw. The findings that transgenic DNA has the tendency to break and
join in several places imply that parts or all of it may be more likely than
the plant's own DNA to jump out of the genome and successfully
transfer horizontally to unrelated species. Horizontal gene transfer, in this
context, means the transfer of the genetic material directly by infection to
the genetic material of unrelated species, in principle to all species
interacting with the GMO: bacteria, fungi, earthworms, nematodes, protozoa,
insects, small mammals and human beings. This process is uncontrollable and
cannot be recalled. The damages done are hence irreversible. Transgenic
DNA has been designed to be invasive and to overcome species barriers;
once released, it will invade different organisms, especially bacteria which
are in all environments, where it will multiply, mutate and recombine.
There are additional findings which suggest an increased potential for the
horizontal spread of transgenic DNA. For example, enzymes that insert the
transgenic DNA into the genome can also help them to jump out again; DNA
released from both dead or live cells can survive without being degraded in
all environments, including the mouth and gut of mammals; DNA can be
readily taken up into cells; and all cells can take up naked or free DNA.
The instability of transgenic DNA may also be enhanced as the result of the
metabolic stress inflicted on the organism by the CaMVpromoter that gives
continuous over-expression of transgenes.

The major consequences of the horizontal transfer of transgenic DNA are the
spread of antibiotic resistance marker genes among bacteria and the
generation of new bacteria and new viruses that cause diseases from the
many bacterial and viral genes used. The generation of new viruses could
occur by recombination with live or dormant viruses that we now know
to be present in all genomes, plants and animals included. Recombination
with defective, dormant animal viral promoters may also occur, as we
know that there are modules within the promoter that are interchangeable
between plant and animal promoters. Recombination of CaMV promoter
modules with defective promoters of animal viruses may result in
recombinant promoters that are active in animal cells, causing over-
expression of one or the other of dozens of cellular genes which are now
believed to be associated with cancer.

There is sufficient scientific evidence to support well-founded suspicion of
serious, irreversible harm to justify the immediate withdrawal of all GM
crops and products containing the CaMV promoter from environmental