Scientific Followup of Dangers of CaMV Virus Used in Genetic Manipulation
Return to
How to Avoid Genetically-Manipulated Soy
Return to
Soy Info Online!
================================
Date Posted: 07/19/1999
Posted by: Dr. Mae-Wan Ho
================================
Here are replies to Roger Morton from Mae-wan Ho and comments from Joe
Cummins, Dept. of Plant Sciences, University of Western Ontario, London
Ontario.
Mae-Wan Ho :
1. 'Naked DNA' is a term which includes DNA outside cells or, in the case of viruses, stripped of the viral coat,
whether they happen to be covered with nucleoproteins or not. See Traavik, T. (1999) Too early may be too late, Ecological risks
associated with the use of naked DNA as a biological tool for research, production and therapy, Research report for Directorate for
Nature Management, Norway.
The CaMV promoter is indeed manipulated in the naked form and incorporated in gene-expression cassettes that are in
turn spliced into the T-DNA of Agrobacterium. The fact that the T-DNA is covered by viral protein is irrelevant,
partcularly as the vir protein is actually needed for conjugation to take place between the Agrobacterium and the plant cell
whereby the T-DNA is introduced into the plant cell genome.
In any case, there is now very good evidence that the CaMV promoter is a recombination hotspot even when it is spliced
into the Agrobacterium T-DNA and introduced into transgenic rice. See Kohli, A. et al (1999). The Plant Journal 17, 591.
2. All naked DNA, in fact, is efficiently taken up by cells of all species including mammals. So much so that naked
DNA in the form of plasmids or other constructs are now used in experiments in gene therapy. There is now an
enormous literature on that subject. Viral and plasmid DNA fed to mice and partially degraded in the gut have been
shown to pass into gut cells, and via the blood stream into blood, liver and spleen cells, where the fragments integrated
into the cellular genome. See Schubbert, R., Lettmann, C. & Doerfler, W. (1994). Ingested foreign (phage M13) DNA survives
transiently in the gastrointestinal tract and enters the bloodstream of mice. Mol. Gen. Genet. 242: 495-504; Schubbert, R., Renz, D.,
Schmitz, B. and Doerfler, W. (1997). Foreign (M13) DNA ingested by mice reaches peripheral leukocytes, spleen and liver via the intestinal
wall mucosa and can be covalently linked to mouse DNA. Proc. Natl. Acad. Sci. USA 94, 961-6.
Another point to consider is that the CaMV promoter in gene-expression cassettes are in unnatural constructs, which
may be more unstable, and hence more recombinogenic than the same in intact viral DNA.
3. The CaMV promoter will be part of the 'naked DNA' of GM plant cells which have disintegrated in the gut. The fact
that it is incorporated into the plant chromosomes is irrelevant, as there is experimental evidence that such DNA released
from GM plants can transform soil bacteria and fungi. See for example, Hoffman, T., Golz, C. & Schieder, O. (1994). Foreign
DNA sequences are received by a wild-type strain of Aspergillus niger after co-culturewith transgenic higher plants. Current Genetics 27:
70-76; Gebhard, F. and Smalla, K. (1998). Transformation of Acinetobacter sp. strain BD413 by transgenic sugar beet DNA. Appl. Environ.
Microbiol. 64, 1550-4; De Vries, J. and Wackernagel, W. (1998). Detection of nptII (kanamycin resistance)
genes in genomes of transgenic plants by marker-rescue transformation. Mol.Gen. Genet. 257, 606-13.
There is no direct experimental evidence yet that transgenic DNA from plants can gain access into bacterial cells or
mammalian cells, but that is because this possibility has never been investigated.
4. Because the CaMV promoter has a recombination hotspot, it is prone to break and join with other DNA. Furthermore,
the T-DNA itself into which the CaMV promoter is spliced has recombinogenic left and right borders showing
substantial analogy to the recombination hotspot in the CaMV promoter. That means the integrated T-DNA could also
break and join with other DNA. That increases the likelihood that the CaMV promoter in DNA of transgenic plant
material taken into cells may become integrated into cellular genomes. Admittedly, the relevant experimental
investigations have not yet been done, but this possibility cannot be dismissed a priori, and should be addressed
appropriately.
Transposable elements are in all genomes, as are relict viral sequences. However, they are generally regulated by
mechanisms not yet fully understood, especially if they have entered the genome a long time ago. Methylation and gene
silencing is one major mechanism for preventing
mobility. However, it is known that stress can induce transpositions, and it is also known that new elements entering the
genome tend to jump around until they become 'tamed'. The integrated transgenic DNA may be similar to new invasive
elements in the genome.
As I have already stated, CaMV promoter within the intact virus will not gain access to mammalian cells because the host
specificity is determined by the viral coat. As mentioned above, the CaMV promoter in gene-expression cassettes are in
unnatural constructs, which makes it more unstable, and hence more recombinogenic than the same in intact viral DNA.
Other comments below from Prof. Joe Cummins, who first drew attention to the dangers of CaMV promoter.
Some points for Morten. CaMV is naked with the gene gun and coated with agrobacterium T system. The CaMV
promoter contains several genes all cis acting, these include the TATA box and the translation enhancer.It is very strange
indeed to proclaim that cis acting genes are not genes at all! The promoter is infectious in the sense that it is
transformable (incorporated as
DNA).The claim that such transformation does not take place is unreasonable. Uptake of DNA into chromosomes after
ingestion has been observed both by Doerfler's laboratory and by experiments reported in human gene therapy. CaMV
does not infect corn and its infection rate is not high even in cauliflower in broccoli. Even infected plants deliver
magnitudes fewer virus sequences than do trangenic plants with CaMV sequence in every cell. The recombinogenic
activity of the CaMV promoter should not be taken lightly nor dismissed with out thought.
http://www.soyinfo.com/haz/ho-camv2.shtml