Proceedings of the XLVII Italian
Society of Agricultural Genetics - SIGA Annual Congress
Verona,
Italy - 24/27 September, 2003
ISBN 88-900622-4-X
Poster
Abstract - 1.14
DIFFERENTIAL GENE EXPRESSION INDUCED BY OZONE
STRESS IN SENSITIVE AND TOLERANT POPULUS SPP.
M. RIZZO*, M.
SALVINI**, R. BERNARDI*, C. NALI***, G. LORENZINI***, M. DURANTE*
*) Dipartimento
di Biologia delle Piante Agrarie, Sezione di Genetica, Via Matteotti 1/B, 56124
Pisa
mrizzo@agr.unipi.it
, rbernard@agr.unipi.it , mdurante@agr.unipi.it
**) Scuola
Normale Superiore di Pisa, Piazza dei Cavalieri 7, 56126 Pisa
***)
Dipartimento di Coltivazione e Difesa delle Specie Legnose "G.
Scaramuzzi", Via del Borghetto 80, 56124 Pisa
cnali@agr.unipi.it , glorenz@agr.unipi.it
ozone stress, Populus, suppression subtractive
hybridisation
Although
definitive proof of a link between ozone and forest damage exist only in
California, large number of data supports a role for this tropospheric
pollutant in forest decline in eastern USA and Europe in combination with other
biotic and abiotic factors.
Plant subjected
to elevated (but often close to reality) concentrations of tropospheric ozone
undergo several biochemical changes, before any morphological damage can be
detected, that are the direct result of a qualitative and quantitative
modification of gene expression.
A lot of
regulating events, responsible for specific physiological reactions, often
depend on the activation or inactivation of particular genes: identification of
differentially expressed genes after ozone treatment can help to understand
molecular mechanisms that are at the basis of the response to this kind of
stress.
In order to
identify differentially expressed genes after the exposition to an acute ozone
dose (a single pulse of 150 ppb for five hours) we have used suppression
subtractive hybridisation, SSH in two poplar species characterised by a
different sensibility to this kind of pollutant: Populus deltoides x maximowiczii,
Eridano clone, e Populus x euoramericana,
I-214 clone, sensitive and tolerant to O3 respectively.
This method
allows to obtain cDNA library enriched of transcript only present in one of the
two compared samples (in our case treated sample and control sample).
The main feature
of this technique is the peculiar effectiveness in isolating rarely expressed
transcripts. This minor fraction is particularly of interest, since transcript
for many regulatory proteins fall into this category.
By using SSH,
four cDNA libraries were obtained, two libraries made of products of genes
activated by stress treatment in sensitive and tolerant plants separately (FS
and FT libraries), and two libraries made of genes whose transcription is
suppressed or diminished under ozone stress, once again in sensitive and
tolerant plants separately (RS and RT libraries).
A primary
screening for all libraries was performed, and several differentially expressed
cDNA clones were selected. From FT library e.g. we have isolated phenyalanine
ammonia-lyase (PAL), ascorbate peroxidase (APX), an important antioxidant
enzyme that helps plants to survive oxidative stress and a cDNA sequence that
presents a conserved domain typical of transcription factors belonging to the
WRKY superfamily that include members involved in the regulation of various
physiological program as pathogens defense.
The expression of some of the most interesting isolated clones has been analysed furthermore.