1 INTRODUCTION
Environmental stress represents the most important single factor
limiting crop yield worldwide, a situation that will certainly aggravate
in the near future as a consequence of global climate changes (IPCC
2014; Buchanan, B.B., Gruissem, W. & Jones 2015). Among the adverse
conditions that constrain plant growth and reproduction, drought has the
highest impact in quantitative terms (FAO 2014). Projections of
increased desertification in several Mediterranean-type environments of
Europe, Australia, South America and sub-Saharan Africa are consistent
across simulation models (IPCC 2014). Water restriction negatively
affects photosynthetic rates by decreasing CO2availability as a result of stomatal closure, and by feedback inhibition
due to limitations in photosynthate transport to sink organs (Romero,
Alarcón, Valbuena & Galeano 2017; Urban, Aarrouf & Bidel 2017). Many
changes associated to water deficit are therefore detected in the leaves
and accordingly, drought responses are inextricably linked to
photosynthesis and chloroplast biochemistry (Zingaretti, Inácio, de
Matos Pereira, Paz & de Castro França 2013).
Potato (Solanum tuberosum L.) is the third most important food
crop in the world (Visser et al. 2009; FAO 2014), and is
vulnerable to drought, salinity and other environmental stresses, which
affect tuber yield and quality (Vasquez-Robinet et al. 2008;
Muñiz García, Cortelezzi, Fumagalli & Capiati 2018). The situation is
particularly critical in developing countries where potato is most
important as an affordable and nutritionally rich food supply (Romeroet al. 2017), and where the impact of global climate change is
predicted to be more severe (IPCC 2014; Muñiz García et al.2018). Therefore, breeders face increasing pressure to develop new lines
with improved drought tolerance while keeping high crop yield, tuber
quality and market acceptance (Romero et al. 2017).
Water limitation elicits a very complex plant response, which combines
physiological, cellular and metabolic adaptations to the stress
situation, and involves genome-wide changes in gene expression patterns.
While many drought-responsive genes have been identified, it is
presently difficult to define the role played by most of them in the
tolerance against this environmental challenge (André et al.2009). As most other abiotic stresses, water deficit causes a rise of
reactive oxygen species (ROS) levels, especially in leaves (Gómez,
Vicino, Carrillo & Lodeyro 2019). In turn, the drought response of
genes involved in ROS metabolism depends on the plant cultivar, its
degree of tolerance and the duration and intensity of the stress
treatment (Cruz De Carvalho 2008). Vasquez-Robinet et al. (2008) found
that the higher drought tolerance displayed by Andean potato genotypes
was related to enhanced expression of genes encoding antioxidant
proteins located in chloroplasts. Moreover, potato transformation with
genes related to ROS scavenging led to lines exhibiting improved
performance under water deprivation (Ahmad et al. 2010; Eltayebet al. 2011; Cheng, Deng, Kwak, Chen & Eneji 2013), indicating
that manipulation of ROS metabolism is a promising strategy to improve
drought tolerance (Gómez et al. 2019).
We have generated tobacco plants with increased tolerance to multiple
sources of abiotic stress by introducing a cyanobacterial flavodoxin
(Fld) directed to chloroplasts (Tognetti et al. 2006; Zurbriggenet al. 2008). Fld is an electron carrier flavoprotein present in
bacteria and some algae that displays essentially the same activities
and redox interactions as the iron-sulfur protein ferredoxin (Fd),
including functional integration as a final electron acceptor in the
photosynthetic electron transport chain (PETC). Fld expression is
normally induced in microorganisms under conditions of iron starvation
and environmental stress that cause Fd down-regulation, taking over Fd
functions as its levels decline, and preventing over-reduction of the
PETC and ROS propagation (Zurbriggen et al. 2008; Pierella
Karlusich, Lodeyro & Carrillo 2014). The Fld gene is absent from plant
genomes (Pierella Karlusich, Ceccoli, Graña, Romero & Carrillo 2015),
but introduction of a plastid-targeted Fld improved delivery of reducing
equivalents to productive pathways of the chloroplast, which in turn
restricted plastid ROS production and increased tolerance to drought and
other stresses (Tognetti et al. 2006, 2007; Zurbriggen et
al. 2008; Li et al. 2017).
The need for drought-tolerant potato lines prompted us to evaluate the
Fld approach in this crop. e describe herein the preparation and
characterization of potato plants expressing a plastid-localized Fld,
and displaying improved photosynthesis, growth and tuber yield under
conditions of water deprivation. To gain further insights into the
mechanism(s) of Fld-associated stress tolerance, we generated
genome-wide transcript profiles from wild-type (WT) and Fld-expressing
potato leaves at a pre-symptomatic stage of water restriction to
evaluate early responses to hydric stress, and combined this approach
with metabolic profiling of carbohydrates and amino acids. The results
provide a detailed snapshot of how chloroplast redox biochemistry
affects gene expression metabolism in a major crop during this
agronomically relevant abiotic stress.