Download PROJECT PROPOSAL for applicants for ITC fellowships

PROJECT PROPOSAL
for applicants for ITC fellowships (2017/18
supervisor:
institution:
contact:
CV:
László SZABADOS, Ph.D., D.Sc.
Institute of Plant Biology
[email protected]
http://www.brc.hu/personal_page.php?id=nb_szlá
project title:
CHARACTERIZATION OF STRESS REGULATORY GENES
FROM EXTREMOPHILE PLANTS
PROJECT SUMMARY
Using the Conditional cDNA Overexpressing system, we have identified several cDNA from
Arabidopsis thaliana and the halophytic relative, Lepidium crassifolium, which can modulate
responses to environmental stresses such as drought and salinity. The research program of the ITC
fellow includes the characterization of 1-2 cDNA clones, which, upon overexpression, can increase
tolerance to osmotic and/or salt stress. cDNA clones will be cloned and characterized by molecular
methods. Tolerance of transgenic lines will be evaluated by phenotypic analysis in controlled
conditions, physiological assays and molecular analysis. Bioinformatic analysis will be used to
characterize phylogenetic relationship, structural features and expression profiles of the
characterized proteins and genes.
BACKGROUND
Drought, soil salinity or extreme temperatures
are representing serious limitations for
Agricultural productivity. Research on model
organisms such as Arabidopsis thaliana has
identified a number of genes and regulatory
pathways, which control the networks linking
stress perception and metabolic or
developmental responses (Cramer, et al.,
2011). Study of a stress sensitive model has
however limitations in understanding tolerance
to harsh environments. Extemophile plants,
such as xerophytes and halophytes can grow in
arid regions or on saline soils, which are
otherwise lethal to non-adapted species.
Halophytes represent 1% of all plant species,
can optimally thrive in the presence of 50-250
mM NaCl, some withstand salt concentrations
up to 600 mM NaCl (Flowers and Colmer,
2008). Features which influence tolerance are
uptake, transport and sequestration of toxic
ions (mainly Na+, Cl-), regulation of cytosolic
K+ retention, optimization of water use, control
of stomata aperture, regulation of osmotic
adjustment through osmoprotectants, and
control of oxidative damage through
detoxification of reactive oxygen species (ROS)
(Flowers and Colmer, 2008). While physiology
of halophytes was extensively studied,
molecular regulation of the extremophile
character remains still to be understood.
Extremophile relatives of Arabidopsis posses
different degree of tolerance to salt, drought,
cold, waterlogging or nutrient limitations
(Amtmann, 2009, Orsini, et al., 2010). Genome
sequences of several extemophile species have
been determined, including T. salsuginea and
T. parvula facilitating the identification of genes
implicated in stress tolerance (Dassanayake, et
al., 2011, Wu et al., 2012).
CURRENT RESEARCH
The Szabados lab conducts research on
responses to to salt and osmotic stress using
Arabidopsis thaliana as main model. The
Conditional cDNA Overexpressing System was
developed in the lab facilitating the identification
of several genes from Arabidopsis, which were
able to improve tolerance to salt, osmotic or
hypoxic stress (Papdi et al., 2008, 2009, Rigó et
al., 2012). The zinc finger factor ZFP3 was
found to regulate ABA sensitivity and influence
the expression of a set of stress and lightinduced genes. Overexpression of the heat
shock factor HSFA4A was found to enhance
tolerance to several environmental stresses,
including salinity, heavy metals, oxidative
agents or hypoxia. The AP2 domain
transcription factor controls tolerance to hypoxia
and osmotic stress and modulates sensitivity to
abscisic acid, key hormone of drought tolerance
(Papdi et al., 2015). Recently, the COST
system have been adapted to a halophytic
relative of Arabidopsis, Lepidium crassifolium,
and several genes have been identified, which
could enhance tolerance of this model plant to
salt or oxidative stresses (Rigó et al., 2016).
SPECIFIC AIMS
Characterization of novel genes identified in the
halophitic plant, Lepidium crassifolium which is
responsible fdor tolerance to salt, drought or
other stresses. Particular aims are the
following:
 Characterization of stress tolerance of 1-3
Arabidopsais COS lines, which carry and
overexpress a Lepidium cDNA.
 Cloning the Lepidium cDNA from the
transgenic Arabidopsis plants, molecular
charactzerization of them.
 Study molecular and physiological
characters of the transgenic lines to get
information on the tolerance trait.
METHODS TO BE LEARNED / APPLIED
 Molecular cloning of cDNA, sequence
analysis, vector construction.
 Determination of plant growth in sterile and
greenhouse conditions, image analysis,
fresh and dry weight determination.
 Analysis of physiological characters which
are related to stress responses: proline
accumulation, photosynthetic parameters,
accumulation of reactive oxygen species
(ROS), lipid peroxidation, determination of
antioxidant capacity, enzyme activities, etc.
 Analysis of molecular characters: eg. gene
expression study by RT-PCR.
 Genetic transformation of Arabidopsis and
perhaps other plants.
 Bioinformatics: use of internet-based
databases, sequence homology search,
determination of protein structure, etc.
SUGGESTED READINGS
Ahuja I, et al.: Plant molecular stress responses face
climate change. Trends Plant Sci, 15:664-674(2010)
Amtmann A: Learning from evolution: Thellungiella
generates new knowledge on essential and critical
components of abiotic stress tolerance in plants. Mol
Plant, 2:3-12(2009)
Cramer GR, et al.: Effects of abiotic stress on plants: a
systems biology perspective. BMC Plant Biol,
11:163(2011)
Dassanayake M, et al.: The genome of the extremophile
crucifer Thellungiella parvula. Nat Genet, 43:913918(2011)
Flowers TJ, Colmer TD: Salinity tolerance in halophytes.
New Phytol, 179:945-963(2008)
Orsini F, et al.: A comparative study of salt tolerance
parameters in 11 wild relatives of Arabidopsis
thaliana. J Exp Bot, 61:3787-3798(2010)
Wu HJ, et al.: Insights into salt tolerance from the
genome of Thellungiella salsuginea. Proc Natl Acad
Sci USA, 109:12219-12224(2012)
Different
molecular,
physiological
and
biochemical technologies will be used during
the program:
SNAPSHOTS FROM THE HOST LABORATORY
Significant publications
Joseph MP, et al.: The Arabidopsis ZINC FINGER PROTEIN3 Interferes with Abscisic Acid and Light Signaling in Seed
Germination and Plant Development. Plant Physiol, 165:1203-1220(2014)
Papdi C, et al.: Functional identification of Arabidopsis stress regulatory genes using the controlled cDNA
overexpression system. Plant Physiol, 147:528-542(2008)
Papdi C, et al.: Genetic technologies for the identification of Arabidopsis genes controlling environmental stress
responses. Funct Plant Biol, 36: 696-720(2009)
Supported by the
TÁMOP 4.1.1.C -13/1/KONV.2014-0001
project
Papdi C, et al.: The low oxygen, oxidative and osmotic stress responses synergistically act through the ethylene
response factor VII genes RAP2.12, RAP2.2 and RAP2.3. Plant J, 82:772-784(2015)
Perez-Salamo I, et al.: The heat shock factor A4A confers salt tolerance and is regulated by oxidative stress and the
mitogen-activated protein kinases MPK3 and MPK6. Plant Physiol, 165:319-334(2014)
Rigó G, et al.: Gene mining in halophytes: functional identification of stress tolerance genes in Lepidium crassifolium.
(submitted)
Representative recent research grants
“Role of heat shock transcription factors and MAP kinases in regulation of plant stress responses.” (OTKA, 2014-2018)
“Epigenetic regulation of ABA signal transduction: function of ZFP3 zinc finger factor in regulation of chromatin
remodeling.” (OTKA, 2016-2020)
Some of the latest students in the laboratory
Zsigmond L, Ph.D., 2003-2008
Papdi C, Ph.D., 2005-2010
Rigó G, B.Sc., M.Sc., Ph.D., 2000-2013
Pérez-Salamó I, Ph.D., 2009-2014
Joseph MP, ITC, Ph.D., 2006-2014
Afwa Thameur A, ITC, 2010-2011
Baba I, ITC, Ph.D., 2014-present
Supported by the
TÁMOP 4.1.1.C -13/1/KONV.2014-0001
project