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Checkbiotech: Plants are given an immune system
Posted by: Prof. Dr. M. Raupp (IP Logged)
Date: September 15, 2005 07:16AM ; ;

Antibodies that can control the function of molecules inside of cells are
important in several applications in both human and plant pathology and
physiology. Unfortunately, most antibodies are unable to function properly
in the cell due to its special environment. Researchers have now been able
to evaluate the intracellular performance of antibodies and constructed them
in such a way so that they can thwart off pathogens or help regulate
cellular events, September 2005 by Katharina Schoebi, Checkbiotech.

An emerging field of immunology that is catching the attention of many
people is immunomodulation, the regulation of the body?s immune system and
other cellular events with antibodies. One example of immunomodulation is
using antibodies to interfere with a pathogen, and by so doing, preventing a
human disease. Immunomodulation, however, is not simple.

When dealing with antibodies, it is important to understand that they
consist of constant and variable domains. The variable domains are important
for binding and contain one disulfide bridge that is fundamental for folding
stability and for antigen-binding properties. However, in the environment of
the cytoplasm, the formation of disulfide bonds is prevented. As a
consequence, the amount of antibody production is considerably lowered, or
prevented altogether.

More recently, researchers have come up with a way to overcome the limiting
factors of antibodies. Scientists have developed what are known as
single-chain antibody fragments (scFv, where the variable domains are joined
by a flexible linker peptide), which provide many of the necessary features
to still survive and carry out certain tasks within the cell, being able to
recognize and attach to their targets appropriately.

This use of scFv intrabodies has proven to be a very important technique to
treat human diseases, so why not try them out in plants too to thwart off
pathogens or help regulate cellular events as well. Such were the thoughts
of Dr. Rosella Franconi from the Ente per le Nuove Tecnologie, l?Energia e l
?Ambiente (ENEA), UTS BIOTEC, in Roma, who recently published her work in
the journal Plant Molecular Biology.

Dr. Franconi and her research group wanted to produce scFv in plants that
recognize the cucumber mosaic virus (CMV), thus providing them with one
basic component of an immune system. CMV is of considerable agronomic
importance, because it infects more than 1,000 plant species worldwide,
particularly tomatoes. Some plants are resistant to the virus. Those that
are not, however, such as tomatoes, are destroyed by CMV because at the
moment, there are no crop protection agents that can prevent CMV infection.

In their experiments, the researchers first screened an antibody library to
obtain scFv structures that best recognized CMV. An antibody library is a
sample of billions of different antibodies. Dr. Franconi and her colleagues
searched in a so called ?F8 library?, composed of very stable scFv molecules
that was created by their lab.

After having found the scFv of interest, the researchers introduced its gene
into minitomatoes so that the plants expressed the antibodies. Subsequently,
the plants were infected with CMV.

The results revealed that the antibodies recognized the pathogen and
hindered an infection. A big breakthough for Dr. Fanconi?s laboratory
occurred when one line of transgenic tomato plants was found to be fully
protected from CMV infection, while previous studies described only partial
protection. In these plants, the virus was able to replicate in the
inoculated cells and spread to adjacent cells, but was not able to spread
throughout the plant.

To ensure that it was the scFv intrabody that was protecting the tomatoes
and not another cellular event, Dr. Franconi?s lab infected the enhanced
tomatoes with a different virus called Tombusvirus Tomato Bushy Stunt
(TBSV). Since the infection with TBSV resulted in disease symptoms both in
transgenic and naturally occurring plants, the researchers conclude that the
scFv protected the tomato plants specifically against CMV.

Further studies should reveal how the scFv intrabodies precisely mediate
protection of plants. In addition, future experiments should also reveal
whether the scFv interfere with processes involved in virus movement by
disturbing interactions with the viral RNA, or other virus or plant cellular

Dr. Franconi?s approach, based on a protein-protein interaction, should
guarantee durable resistance against most CMV types, because they all have
an almost identical structure that is recognized by antibodies. Since this
structure has remained intact as CMV has evolved, there are only slight
chances that CMV would become resistant to Dr. Franconi?s scFv intrabodies.

Dr. Franconi sees her enhanced tomato varieties someday benefiting the
Mediterranean area, which is a strong producer of tomatoes. If concerns were
to arise about spread to wild varieties, Dr. Franconi told Checkbiotech,
?Moreover, the tomato is a predominantly self-pollinating plant and in the
Mediterranean area there are no wild tomato species that are sexually
compatible with the cultivated ones.?

Dr. Franconi?s work provides the first evidence that the ?F8 library? ?
developed at ENEA ? contains intrabodies that are able to modulate
biological processes in living cells. Furthermore, it offers the opportunity
to easily isolate stable molecules against several plant and animal targets
and thereby overcomes the limitations of current immunomodulation

The researchers have done their studies with CMV and tomato and are now
interested in carrying out similar studies with other viruses, as well as
with human-related diseases (virus, oncogenes, etc.). However, this kind of
research is very underfunded, Dr. Franconi said. ?At present, we do not have
financial support to carry on with such studies.? Her work has great
potential to eliminate the yearly losses caused by CMV in tomato crops and
others that occur globally.


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