The need for aquatic model crops is only exacerbated by increasing market value in indoor hydroponic cultivation systems.Root structural architecture defines the spatial configuration of the root system and variation in RSA can reflect efficient phosphate uptake by plants.Much of the current literature focuses on RSA traits in maize , common bean and Arabidopsis thaliana with RSA shown to be a highly plastic trait, changing in response to the availability of water, nutrients and hormone signalling. For example, ethylene is involved in the promotion of lateral root growth, root hair growth and inhibition of primary root growth under low P conditions. Reduced root growth angle : is one of the most important RSA traits for improving P acquisition in many soil-grown species. In maize and common bean, lower RGA is associated with increased P accumulation and improved growth in P deficient soil where P is concentrated in the topsoil. However, in aquatic systems when P is likely homogenously distributed, a lower RGA is unlikely to be advantageous. Root growth angle therefore is not considered important as a trait for enhance PUE in aquatic plants. Increased lateral root density:also enhances P acquisition by allowing plants to explore outside P-depleted zones. Using maize recombinant inbred lines with contrasting lateral rooting phenotypes, significant differences in phosphorus acquisition, biomass accumulation and relative growth rate were observed under low phosphorus availability. Increased investment in the production of lateral roots was shown to be cost-effective under low P. For aquatic crops,nursery pot enhanced lateral root density is an important trait for enhanced PUE since it increases root surface area for P uptake. Adventitious roots: from above ground structures can enhance topsoil foraging by up to 10% in stratified soil.
These require a lower metabolic investment than basal roots, however, in uniform soil they can limit P acquisition by hindering the growth of basal roots. In aquatic and f looding-tolerant plants, adventitious roots are important for P uptake within the water column. There is positive correlation between the size of the adventitious root system and P uptake in bittersweet plants under long-term submergence, and as mentioned, adventitious roots are responsible for a higher proportion of P acquisition in watercress than basal roots. Thus, in an aquatic crop such as watercress, increased adventitious root production is a key trait for enhanced PUE.Modelling of root architecture in maize and bean has shown RCA may increase the growth of plants up to 70% in maize and 14% in bean under low P availability largely due to remobilisation of P from dying cells. However, formation of aerenchyma is also associated with reduced root hydraulic conductivity in maize which may impede transport of water, but is unlikely to be relevant to hydroponically grown crops. Most aquatic plants, including watercress, form aerenchyma constitutively in roots and stems to aid internal gas exchange and maintain strength under water pressure. For aquatic crops such as watercress, formation of root aerenchyma is an important trait for selection for enhanced PUE. Root hair density and root hair length: root hair density increases up to 5 times in low P conditions. Using Arabidopsis mutants, Bates & Lynch found hairless plants had lower biomass and produced less seed than wild-type plants at low phosphorus availability. Root hairs increased root surface area by 2.5 to 3.5-fold in barley and wheat , respectively, and there was an almost perfect correlation between P uptake and root hair surface area. Root hair traits vary substantially between genotypes and the genetic control underlying their formation is well understood, thus making them an excellent target for plant breeding programs . Root hair length and density are likely to be important for PUE in aquatic crops as they significantly increase root surface area for P uptake.A modelling approach in Arabidopsis showed that the combined effects of root hair length, root hair density, tip to first root hair distance and number of trichoblast files on P acquisition was 3.7-fold greater than their additive effects. For aquatic species such as watercress, the root ideotype for determining optimal P acquisition remains unknown.
Although, absorption of P through the shoots is still debated, root uptake is generally regarded as the mode of P uptake in aquatic plants. Watercress beds have a fine gravel substrate which contains negligible amounts of P. The substrate within watercress beds is likely too shallow to allow for significant stratification of P, thus a shallower basal root angle would be unlikely to provide much adaptive benefit. In groundwater sources, P may be distributed more homogenously due to turbulent f lowing water, so RGA will not assist within the water column. Nevertheless, even with homogenous P distribution, plants with shallower root systems have been shown to encounter less inter-root competition with roots on the same plant so RGA could provide an adaptive value in this sense. Cumbus & Robinson studied P absorption by the adventitious and basal roots of watercress and found that the adventitious roots absorbed a higher proportion of P at low P concentrations, despite having a lower biomass compared to the basal root tissue. Thus, adventitious roots are also a key trait for analysing watercress PUE. Increased production of lateral roots, adventitious roots and root hairs all increase root surface area, and thus will increase P acquisition from the water and sediment and are important traits for PUE in watercress. In addition, the root cap can account for 20% of the phosphate absorbed by the roots of Arabidopsis. Therefore, increasing the number of roots increases the number of root tips and the number of these “hot spots” for phosphate acquisition.Plants are reliant on phosphate transporters to acquire P from the environment and transport P between tissues, and this includes for aquatic plants. The PHT1 family is the most widely studied group of P transporters and is primarily responsible for P uptake but also has a role for P transport between tissues. A broad range of expression patterns are associated with different PHT1 genes but generally, higher expression of PHT1 genes is associated with improved shoot biomass accumulation and P tolerance. Watercress with higher PHT1 expression may result in improved biomass accumulation in P deficient water, but this has yet to be tested. Additional traits that are important in other crops are organic acid exudation and phosphatase activity. Since these control release of P from organic forms in the soil, they are less relevant to watercress cultivation where P released from bound sources would be rapidly lost to the watercourse.
However, phosphatases that remobilise P from intracellular sources have been identified in Arabidopsis so similar phosphatases could enhance internal P utilisation in watercress.Alongside phosphate acquisition, PUE also refers to more efficient P utilisation associated with re-translocation and recycling of stored P, that relies on effective P transportation within the plant, P scavenging, and use of alternate biochemical pathways that bypass P use. Re-translocation between plant tissues is governed by transporters such as PHT transporters and PHO transporters. Unlike, PHT transporters which regulate P acquisition too, PHO transporters are solely responsible for P transport into vascular tissues and cells. The genetic control underlying these PUE mechanisms is covered in the subsequent section. Alternative P use strategies includes substituting phospholipids in cell walls with sulfolipids and galactolipids. Several enzymes in the glycolytic pathway depend on P so bypass enzymes such as pyrophosphate dependent phosphofructokinase , large pots plastic phosphoenolpyruvate carboxylase and pyruvate phosphate dikinase can be recruited to use pyrophosphate for a P donor and conserve limited ATP pools. Several studies have reported increased PEPC activity under P deprivation. The mitochrondrial electron transport chain responds by utilising non-phosphorylative pathways. Acid phosphatases in intracellular spaces or present in the apoplast can increase P availability by remobilising P from senescent tissues and the extracellular matrix. Both aspects of PUE rely on accurate sensing of the P state within the plant and external environment to alter global gene expression and ensure appropriate responses to upregulate P uptake and P use pathways.QTL for overall PUE metrics as well as QTL for more specific architectural root traits associated with low P tolerance have been identified in several economically important crops including soybean, soybean , rice , maize and common bean. RSA is extremely plastic, subject to effects of hormone signalling, environmental stimuli and under the control of several genes so elucidating these QTL is challenging. Studies on other Brassicaceae species are likely of most genetic relevance for QTL mapping in watercress, however QTL associated with other species such as soybean, rice, sorghum and wheat are summarised in Table 1. P-starved Arabidopsis exhibit longer root hairs and higher root hair density, decreased primary root length and increased lateral root density. Three QTL, were identified which explained 52% of the variance in primary root length. In rapeseed primary root length decreases, lateral root length and density increases with declining P concentration. Several QTL are associated with these changes and many co-locate with QTL for root traits in Arabidopsis. A more recent study used over 13 000 SNP markers to construct a genetic linkage map in rapeseed, where 131 QTL were identified in total across different growth systems and P availabilities. However, only four QTL were common to all conditions, demonstrating strong environmental effects determining these QTL. To date, there is no published literature on QTL associated with aerenchyma formation under low P in any plant species and no studies exist on QTL mapping for root traits in watercress. Identification of QTL and markers associated with PUE could accelerate breeding for nutrient use and reduce the environmental impact associated with watercress cultivation.
Specific genes involved in root architecture are targets for enhanced PUE. Although RSA traits are highly quantitative, a BLAST to the rapeseed reference genome revealed 19 candidate genes related to root growth and genetic responses to low P in Arabidopsis. These genes included AUXIN-INDUCED IN ROOT CULTURES 12involved in auxin-induced production of lateral roots and PHOSPHATE DEFICICENCY RESPONSE 2which is part of growth changes in the plant apical meristem under P deficiency. PDR2 is a major component of the P starvation response and functions togetherwith LPR1 and its close paralog LPR2 as a P-sensitive checkpoint in root development by monitoring environmental P concentration, altering meristematic activity and adjusting RSA. Genes involved in transcriptional control are multifunctional under P deprivation; some have overlapping roles in RSA development, P signalling and P utilisation. They are discussed together here despite partial involvement in P utilisation. PHR1and PHL1code for transcription factors that play critical roles in the control of P starvation responses. PHR1 mediates expression of the microRNA miR399 which modulates the PHO2 gene, responsible for P allocation between roots and shoots and affects expression of other PSR genes such as PHT transporters. SPX transcription factors are important negative regulators of PSR via repression of PHR. The roles of several other transcription factor genes on RSA and other regulatory elements are summarised in Table 2 and Figure 3. Auxin, sugars and other hormones such as cytokinins, ethylene, abscisic acid , giberellins and strigolactones are implicated in phosphate-induced determination of RSA so genes involved in these pathways may be significant candidates. Under low P, auxin levels increase in root hair zones and root tips. Auxin mutants such as taa1and aux1have impaired root hair growth in low P. Expression of the Arabidopsis auxin receptor gene TIR1 increases under low P availability which results in increased sensitivity to auxin and production of lateral roots.Mutants in auxin-inducible transcription factors also have disrupted root hair responses under low P. ROOT HAIR DEFECTIVE 6- LIKE-2and ROOT HAIR DEFECTIVE 6-LIKE-4are responsive to P deficiency and promote root hair initiation and elongation. ARF19 is a key transcription factor promoting auxin-dependent root hair elongation in response to low P. HPS1is involved in regulating the sucrose transporter SUC2 and hps1 mutants exhibit significant P-starvation responses under P-sufficient conditions. Plants with impaired cytokinin receptors CRE1 and AHK3 show increased sugar sensitivity and increased expression of P-starvation genes. ETHYLENE RESPONSE FACTOR070 is a transcription factor critical for root development under P starvation. Though no studies exist for P-associated gene expression changes in watercress, Müller et al. used RNA sequencing approaches to identify responses to submergence in watercress and found several ABA biosynthesis and catabolism genes associated with stem elongation. This study provides a model for using transcriptomic approaches to explore hormone-induced morphological changes in watercress.