Introduction
Limited water availability is a shared component of drought and salinity stresses that constrains crop growth and yield. Additionally, salinity stress limits plant growth and agricultural productivity through nutritional imbalance and ion toxicity. Roots sense their environment, triggering transcriptomic and biochemical responses that allow the plant to adapt to such conditions through local and systemic responses, with hormones playing a key role in such adaptive responses (Achard et al. 2006). Root-targeted alteration of hormone metabolism and signalling has been proposed as a biotechnological strategy to overcome the effects of saline soils, and to enable this we must understand the specific adaptive roles of plant hormones (Ghanem et al. 2011b; Albacete, Martínez-Andújar & Pérez-Alfocea 2014).
Crops dynamically regulate their root system architecture (RSA) in response to environmental stresses to fulfil their mineral and water requirements. In dry and saline soils, plants reduce lateral root initiation and elongation while promoting root hair density and the growth of the primary root to reach deeper water and nutrient sources (Xiong, Wang, Mao & Koczan 2006; Ma et al. 2010; Brown et al. 2012; Xu et al. 2013). Depending on the level of salt tolerance of the plant species or genotype, low-moderate salinity (2‑8 dS m-1) can promote root growth while high salt levels (8-16 dS m-1) restrict root development (Julkowska & Testerink 2015).
Among the different plant hormones, tissue-specific ABA levels (and responses) change dynamically according to developmental and environmental stimuli. Although ABA is generally considered to inhibit growth of well-watered plants, low ABA concentrations (< 1 μM) can stimulate root growth of Arabidopsis (Ephritikhine, Fellner, Vannini, Lapous & Barbier-Brygoo 1999; Fujii, Verslues & Zhu 2007). Moreover, wild-type (WT) ABA levels are necessary to sustain root growth in maize seedlings grown under low water potential (Sharp & LeNoble 2002), and for leaf expansion and shoot development in tomato (Sharp, LeNoble, Else, Thorne & Gherardi 2000) and Arabidopsis (LeNoble, Spollen & Sharp 2004) under well-watered conditions. ABA may stimulate growth by restricting the biosynthesis of ethylene, a growth inhibitor (reviewed in Sharp et al., 2004). Within the roots, ABA alters gene expression that induces changes in RSA (Sharp et al. 2004), increases root hydraulic conductivity (Thompson et al. 2007a), modifies nutrient and ionic transport and changes primary metabolism leading to osmotic adjustment (Sharp & LeNoble 2002; Martínez-Andújaret al. 2020b).
ABA is seemingly exported to the shoot as a root-to-shoot signal, since plants growing in dry or saline soil can show stomatal closure before shoot water status (the trigger for ABA accumulation) begins to decline (Gowing, Jones & Davies 1993; Dodd 2005). Stress-induced increases in xylem sap ABA concentration are independent of shoot water status and often inversely correlated with stomatal conductance (gs ) (Wilkinson & Davies 2002). However, experiments with reciprocal grafts of ABA-deficient and WT plants showed that stomatal closure of WT scions in response to dry (Holbrook 2002) or saline (Li, de Ollas & Dodd 2018) soil was rootstock independent. Instead, roots in drying soil alkalise xylem sap causing a redistribution of existing pools of ABA within the leaf that affects stomatal closure (Wilkinson, Corlett, Oger & Davies 1998), and other non-ABA chemical signals such as sulphate (Malcheska et al. 2017) or jasmonic acid (De Ollas, Arbona, Gómez-Cadenas & Dodd 2018) may also be involved. ABA detected in the root system may either be synthesized locally or translocated from the shoot via the phloem (McAdam, Brodribb & Ross 2016), and ABA can recirculate between roots and shoots, with roots either acting as a sink for ABA or as a net exporter of ABA to the shoot, depending on plant nutrient and water status (Peuke 2016).
Genetically increasing endogenous ABA levels is a promising strategy to improve resistance to abiotic stresses such as drought and salinity. The enzyme 9-cis -epoxycarotenoid dioxygenase (NCED) is rate-limiting for ABA biosynthesis, and over-expression of NCED genes increased ABA content of tissues, as first shown in tobacco and tomato by overexpressing the tomato gene SlNCED (Thompson et al.2000, 2007a b). This work provided transgenic tomato lines with different levels of expression of SlNCED1 and ABA contents (SP12 and SP5) and offers the opportunity to study the effects of high ABA on root-to-shoot communication. In previous reciprocal grafting experiments between WT, SP12 and SP5, ABA in xylem sap collected from detopped roots was mainly determined by the root genotype, as might be expected in the absence of the shoot. Also, root cultures (again independent of the shoot) of SP12 and SP5 had higher ABA content that WT, thus overexpression of SlNCED1 was sufficient to increase ABA biosynthesis in the root alone (Thompson et al. 2007b), despite the much lower level of NCED substrate available in roots compared to leaves (Taylor, Sonneveld, Bugg & Thompson 2005). In contrast, stomatal conductance in well-watered reciprocal grafting experiments was significantly affected only by the shoot genotype (Thompson et al. 2007b). Overexpression of NCED has now been explored in many systems, and its limiting effect on stomatal conductance confers improved water use efficiency (WUE) (Thompson et al. 2007a) and resistance to terminal drought (withdrawal of irrigation in pot experiments). The latter effect, e.g. in tobacco (Qin & Zeevaart 2002), grapevine (He et al. 2018), and petunia (Estrada-Melo, Ma, Reid & Jiang 2015) is dominated by the lower transpiration rate and slower soil moisture depletion. NCED overexpression also increased growth relative to WT under osmotic stress (NaCl, mannitol) in tobacco (Zhang, Yang, Lu, Cai & Guo 2008) and improved transpiration and reduced chloride accumulation in Arabidopsis grown in “a 150 mM chloride dominant solution” (Zhang, Yang, You, Fan & Ran 2015). However, the effect of rootstocks overexpressing NCED on plant growth and yield responses to saline soil has not been investigated.
ABA interacts with other hormones to mediate local and systemic stress responses (Sah, Reddy & Li 2016): it antagonizes the growth inhibitory effects of ethylene production in tomato shoots (Sharp et al.2000), Arabidopsis shoots (LeNoble et al. 2004), and maize roots (Spollen, Lenoble, Samuels, Bernstein & Sharp 2000), and also during grain-filling in wheat (Yang, Zhang, Liu, Wang & Liu 2006). Moreover, root-supplied ABA from WT rootstocks was sufficient to revert xylem ACC concentrations, foliar ethylene production and leaf area of ABA-deficient scions (Dodd, Theobald, Richer & Davies 2009). However, night-time maize leaf expansion of water-stressed plants did not appear to be regulated by either ABA or ethylene (Voisin et al. 2006), but probably by more complex hormone interactions.
Many hormones (ABA, ethylene, JA and brassinosteroids) modify the development of RSA in saline stress conditions (Achard et al.2006; Osmont, Sibout & Hardtke 2007; Zolla, Heimer & Barak 2010; Duanet al. 2013; Geng et al. 2013). The integration of auxin and cytokinin antagonistic mechanisms might be mediated by gibberellins, because auxin induces degradation of DELLA proteins and enhances cell cycle activity, whereas gibberellins limit the growth inhibition mediated through cytokinin (reviewed in Petricka et al., 2012). Although salinity leads to root, xylem and leaf ABA accumulation in tomato (Albacete, Martínez-Andújar, Pascual, Acosta & Pérez-Alfocea 2008b; Liet al. 2018), it is not clear whether it directly controls plant responses, since other hormonal factors (such as ethylene precursor ACC and the ratio ACC/ABA) co-varied with the productivity (biomass), photosynthetic parameters and WUE (Cantero-Navarro et al. 2016). These two root-derived hormones were positively (ABA) or negatively (ACC) correlated with productivity in a salinized population of plants in which a common scion was grafted onto rootstocks representing a recombinant inbred line population from the cross S. lycopersicum× S. cheesmaniae (Albacete et al. 2009).
Grafting is a common commercial practice in many woody and herbaceous horticultural species (Albacete et al. 2014), and easily applied in the field. Tomato is one of the most important economic crops in the world and it is commonly propagated by grafting high productivity scions onto vigorous rootstocks to alleviate soilborne diseases and abiotic stress effects (Bletsos & Olympios 2008; Martínez-Andújar, Albacete & Pérez-Alfocea 2020a). Cultivated tomato is moderately tolerant to salinity with a threshold of tolerance of 2.5 dS m-1but there is a subsequent yield loss of 10% for each unit of salinity increase (François & Maas 1994), which means that 30-40% yield losses due to salinity are quite common in many horticultural areas such as the tomato-producing region of Southeast Spain. Root-specific traits such as RSA, sensing of edaphic stress and root-to-shoot communication can be exploited to improve resource (water and nutrients) capture and plant development under resource-limited conditions. Root system engineering and rootstock breeding provides new opportunities to maintain sustainable crop production under changing environmental conditions. We hypothesises that grafting a commercial tomato cultivar scion onto ABA over-producing tomato rootstocks would enhance growth and yield under saline conditions, potentially through multiple local and systemic mechanisms.