[SANET-MG] preventing flowering in trees

[jcummins <jcummins@UWO.CA>]

February 21, 2005
“Preventing flowering in transgenic trees”
               Transgenic(genetically modified , GM) trees are beginning to  flood
the ecosystems of the world. The modified trees have been tested
extensively in large open plots with little concern over the spread of
genes from the plots. Dispersal of pollen and seeds  from forest trees
has been extensively investigated. Reviews of such studies show that
gene flow transport may be measured in kilometers (1,2). It is very
clear that the transgenes from GM trees cannot be contained on a
plantation and they will spread widely. For that reason  a great deal of
effort has been devoted to developing  molecular  genetic modifications
that will prevent flowering or pollen production. Unless fool proof
methods for controlling flower or pollen production are proven
commercial development of transgenic trees is out of the question.
For the most part, the methods  used to control flowering or pollination
involved interfering with the genetic program for floral development or
 for deleting  cells involved in floral development, Before  reviewing
the  various  flower control projects for forest trees it might be
helpful to  briefly examine the genes controlling flowering. A group of
 genes called MADS-box  genes code for transcription factors that
recognize DNA binding domains  (the MADS-box) and recognize similar
target DNA sequences  that determine developmental pathways. The MADS
genes associated with flowering have to be identified  then modified to
control flowering or pollination. The plant MADS genes are related to
animal homeotic (HOX) genes  that are extensively studied regulators of
morphogenesis(3).  Unraveling the functions of MADS genes has allowed
flower development to be manipulated.
Flowering is prevented by  either by  controlling gene product formation
 using anti-sense genes or by using small regulatory RNA to prevent
active gene products such as the MADS box  transcription factor from
being formed or by  genetic ablation using a toxin gene driven by  a
floral gene promoter.. The preferred  toxin gene to be employed is the
barnase ribonuclease from the bacterium  Bacillus amylolquefaciens. That
ribonuclease is activated using a promoter specific to floral or pollen
development. The gene effectively kills  the cells that  it targets.
Another cell death toxin used is the diphtheria  toxin from the
bacterium Cornyebacterium diphtheria  or related  ADP-Ribosyltransferase
toxins form other bacteria but those toxin gene are used less than the
barnase gene. The preferred barnase gene  is a part of  the genetic
construction called “terminator” that has been developed to control
corporate  seed production (5).
Professor Steven  Strauss  of Oregon State University pioneered flower
and pollen control  in poplar.  He and his colleagues have led in the
area of flowering control in forest trees. Strauss pointed out that
when completer floral sterility is achieved  will require vegetative
propagation (6). Floral sterility has begun to  be extended from poplar
to shade trees (7). Strauss has argued that management of GM poplar is
comparable to conventional poplar even though he is well aware of the
seed and pollen dispersal  with transgenic poplar (8). Along with the
exploration  of floral sterility  Strauss has investigated  speeding
flower development (trees normally take years to develop sexually) to
allow rapid breeding  and selection cycles.(9). Of course the rapid
breeding cycle  is fraught with uncertainty  about the mature tree.
Strauss has introduced use of the poplar homologue to the floral MADS
box genes , the poplar promoter gene PTD(10). The PTD promoter was
combined with the diphtheria  toxin gene, DTA,  to produce sterile polar
without the detrimental  effects on yield encountered earlier (11). The
problem of somaclonal variation is hardly mentioned in the discussion of
flower control in poplar even though the problem was discussed in a
report on a four year field trial of herbicide tolerant poplar reported
by the Strauss group (12). Somaclonal variation results from the cell
culture technique used to select and propagate transgenic plants.  It
results in extremely high levels of mutation and chromosome instability
which could reverse floral sterility. Earlier reports showed that
poplar cell culture resulted in extremely high levels of somaclonal
variation(13,14).
In Finland investigators from Sopanen University have  studied the
control of flowering in silver birch. Those investigators identified the
MADS box genes controlling flowering in the birch tree (15,16). When a
flower specific birch promoter gene BpMADS1 was used to drive the
barnase gene  floral cell ablation prevented flowering but there were
marked side effects effecting leaves  and branching (17). The side
effects were likely a pleiotrophic effect of the gene insertion but
could, as well have been effected by somaclonal variation from cell
culture.  A recent report altered the name of the MADS box gene from
BpMADS to BpFULL1 as in the previous study  flowering was prevented but
the gene modification effected leaves and branching (18). The
pleiotrophic effects  observed may extend into  areas not yet detected
and they require more extensive study.
       Before leaving the discussion of  the control of flowering in trees it
may be useful to consider the  potential side effects of the ablation
toxins used to create sterile trees. Barnase ribonuclease proved  toxic
to the kidneys of  rats (19)  Barnase was cytotoxic in  mice and in
human cell lines  (20). Animals may not find the GM forests welcoming.
Diphtheria toxin has been associated with anaphylactic response (221)
As the song goes: “If you go down in the (transgenic) woods today,
You're sure of a big surprise.” The safety of transgenic forest should
be carefully evaluated before  the trees  are widely dispersed.
References
1.DiFazio,S,Slavov,G,Burczyk,J,Leonardi,S. and Strauss,S. Gene flow from
tree plantations and implications for transgenic risk assessment 2004
Plantation Forest Biotechnology for the 21st Century eds. Walter,C. and
Carson,M. 405-22
2. Slavov,G,DiFazio,S. and Strauss,S. Gene flow from forest trees: from
empirical estimates to transgenic risk assessment 2002 Scientific
methods workshop: ecological and agronomic consequences of gene flow
from transgenic crops to wild relatives Columbus,Ohio
3.Ng,M. and yanofsky,M.  Function and evolution of the plant MADS –Box
gen family 2001 Nature Reviews Genetics 2,186-96
4.Skinner,J,Meilan,M, Brunner,A. and Strauss,S.  Options for genetic
engineering of floral sterility in forest trees 2000 Jain,S. and
Minocha,S. eds Molecular Biology of Woody Plants  Kluwer Academic
publishers Netherlands
5.Ho,M. and Cummins,J. 2001 Terminate the terminators!
 http;//www.i-sis.org.uk/
6Strauss,S,Rottman,W,Brunner,A. and Sheppard,L. Genetic engineering of
reproductive sterility in forest trees 1995 Molecular Breeding 1, 5-26
7.Brunner,A,Mphamed,R,Meilan,R,Sheppard,L, Rottman,W. and Strauss,S.
Genetic engineering of sexual sterility in shade trees 1998 J.
Arboculture 24, 263-73
8. Strauss,S.,DiFazio,S. and Meilan,R. Genetically modified poplars in
context 2001 The Forestry Chronicle  77,271-80
9. Strauss, S., and Brunner, A.  Tree biotechnology in the 21st century:
Transforming trees in the light of comparative genomics.(2004). In S.H.
Strauss and H.D. Bradshaw (Eds.), The BioEngineered Forest: Challenges
to Science and Society, Pp 76-97. Resources for the Future, Washington,
D.C., USA.
10. Meilan,R.Brunner,A,Skinner,J. and Strauss,S. Modification of
flowering transgenic trees 2001 eds. Morohoshi,M. and Komamine,A. eds
Molecualr Breeding of Woody Plants  Elsevier Science B.V. 247-57
11. Skinner,J,Meilan,R,Ma,C. and Strauss,S. The poplar PTD promoter
imparts floral predominant expression and enables high levels of floral
organ ablation in Populus, Nicotiana and Arabidopsis” 2003 Molecular
Breeding 12,119-32
12. Meilan,R,Auerbach,D,Ma,C.,DiFazio,S. and Strauss,S. Stability of
herbicide resistance and GUS expression in transgenic hybrid poplars
during four years of ffield trials and vegetative propagation  Hort.Sci.
37,277-80
13. Antoneti,L. and Pinon,J.  Somaclonal variation within poplar 1993 Plant
Cell ,Tissue and Organ culture 35,99-106
14. Wang, G., Castiglione, S., Chen, Y., Li, L., Han, Y. and Tian, Y.
Poplar (Populus nigra L.) plants transformed with a Bacillus
thuringiensis toxin gene:
insecticidal activity and genomic analysis 1996 Transgenic Res. 5, 289–301
15. Lemmetyinen,J,Hassinen,M,Elo,A,Porali,I,Keinonen,K,Makela,H. and
Sopanen,T. Functional characterization of SEPALLATA3 and AGAMOUS
orthologues in silver birch 2004 Physiologia Plantarum 121,149-62
16. Lannenpa,M,Janonen,I,Holtta-Vuor,M,Gardemeister,M,Porali,I. and
Sopanen,T.  A new SBP-box gene BpSPL1 in silver birch (Betula pendula)
2004 Physiologia Plantarum 120,491-500
17. Lemmetyinen,J,Keinonen,K. and Sopanen,T. Prevention of the flowering
of a tree, silver birch 2004 Molecular Breeding 13,243-9
18. Lännenpää,M,Ranki,A,Hölttä-Vuori,M,Lemmetyinen,J,Keinonen,K. and
Sopanen,T. Prevention of flower development in birch and other plants
using a pFULL1::BARNASE construct  2005 Plant Cell Reports in press  doi
10.1007/s00299-004-0903-y
19. Ilinskaya,O. and Vamvakas,S.  Nepherotoxic effects of bacterial
ribonuclease in the isolated perfused rat kidney  1997 Toxicology 120,55-63
20. Prior,T, Kunwar,S. and Pastan,I.  Studies on the activity of barnase
toxins in vitro and in vivo 1996 Bioconjugate Chemistry  7,23-9
21.Rosenberg,A. Immunogenicity of biological therapeutics: a hierarchy
of concerns 2003 Developmental Biology 112,15-21





********************************************************
To unsubscribe from SANET-MG:
1- Visit http://lists.sare.org/archives/sanet-mg.html to unsubscribe or;
2- Send a message to <listserv@sare.org> from the address subscribed to the list. Type "unsubscribe sanet-mg" in the body of the message.

Visit the SANET-MG archives at: http://lists.sare.org/archives/sanet-mg.html
For more information on grants and other resources available through the SARE program, please visit http://www.sare.org.