Mark Dempsey
Dil Thavarajah*- Plant and Environmental Sciences, Pulse Quality and Nutritional Breeding, Biosystems Research Complex, Clemson University, Clemson, SC, United States
Lentil (Lens culinaris Medikus) is a nutrient-rich, cool-season food legume that is high in protein, prebiotic carbohydrates, vitamins, and minerals. It is a staple food in many parts of the world, but crop performance is threatened by climate change, where increased temperatures and less predictable precipitation can reduce yield and nutritional quality. One mechanism that many plant species use to mitigate heat and drought stress is the production of disaccharides, oligosaccharides and sugar alcohols, collectively referred to as low molecular weight carbohydrates (LMWCs). Recent evidence indicates that lentil may also employ this mechanism – especially raffinose family oligosaccharides and sugar alcohols – and that these may be suitable targets for genomic-assisted breeding to improve crop tolerance to heat and drought stress. While the genes responsible for LMWC biosynthesis in lentil have not been fully elucidated, single nucleotide polymorphisms and putative genes underlying biosynthesis of LMWCs have been identified. Yet, more work is needed to confirm gene identity, function, and response to abiotic stress. This review i) summarizes the diverse evidence for how LMWCs are utilized to improve abiotic stress tolerance, ii) highlights current knowledge of genes that control LMWC biosynthesis in lentil, and iii) explores how LMWCs can be targeted using diverse genomic resources and markers to accelerate lentil breeding efforts for improved stress tolerance.
1 Introduction
Lentil (Lens culinaris Medikus) is a nutrient-rich, cool-season food legume produced in dry regions worldwide. Lentil has been cultivated for over 10,000 years and was domesticated in the fertile crescent (Cubero, 1981). Lentil is an herbaceous, self-pollinating diploid with seven chromosomes (2n = 14). Total global production has been increasing over the last two decades (Kaale et al., 2023), with recent production at 6.16 ± 0.47 million tons annually (5-year mean ± standard deviation: 2018-2022), concentrated in Canada (36% of global production), India (22%), Australia (11%), Turkey (6%), and the United States (4%) (FAO, 2024). Production is expected to continue rising in the coming decades to help feed the growing human population, as lentil is high in protein (20 – 25%), rich in carbohydrates (60 – 63%) and many micronutrients, and low in fat (1.5 – 3%) (Johnson et al., 2020; Sen Gupta et al., 2013; Tahir et al., 2011; Thavarajah et al., 2011; Wang and Daun, 2006; Zhang et al., 2014). Lentil seeds contain high concentrations of prebiotic carbohydrates (11 – 25%), which are not directly digested by humans but are fermented in the gastrointestinal tract by beneficial microorganisms, and are associated with a healthy gut microbiome and other health benefits (Beserra et al., 2015; Gibson et al., 2017; Johnson et al., 2021). Lentil prebiotic carbohydrates consist of three basic carbohydrate classes: oligosaccharides (0.9 – 6.1 g/100 g), sugar alcohols (SAs; 0.25 – 1.7 g/100 g), and resistant starch (3.7 – 22.1 g/100 g) (Johnson et al., 2015, 2021). There are two classes of oligosaccharides in lentil: raffinose family oligosaccharides (RFOs; 0.9 – 6.0 g/100 g) and fructooligosaccharides (FOS; 0.06 – 0.09 g/100 g), with RFOs constituting the vast majority (ca. 97 – 99%) of oligosaccharides in lentil (Table 1).
Table 1. Concentration ranges (mg/100 g) of LMWCs and broad sense heritability estimates (H2) in pulse crops.
Oligosaccharides and SAs are critical throughout the lifecycle of many plant species, including legumes, as they improve tolerance to abiotic stress, such as high temperatures, drought, saline conditions, and oxidative stress (Benkeblia, 2022; Merchant and Richter, 2011; Yan et al., 2022). Sucrose and other disaccharides also improve tolerance to abiotic stress (Guy et al., 1992; Koster, 1991; Morelli et al., 2003). RFOs, FOS, SAs, and mono- and disaccharides are referred to in this review article as low molecular weight carbohydrates (LMWCs) and are defined in the following section. While the role of LMWCs in stress tolerance has not been explicitly studied in lentil, recent research showed LMWCs in lentil seeds varied significantly across nine environments, with the total LMWC concentration positively correlated with growing season temperature (Johnson et al., 2015). This research suggests that higher temperatures can lead to more LMWC accumulation in seeds, possibly in response to heat or water-deficit stress. Thus, because of its favorable nutrient profile and potential stress tolerance, lentil is a good candidate for adaptation to a changing climate.
Higher temperatures and frequent droughts will put unprecedented pressure on crop production systems within the next century (Coughlan de Perez et al., 2023; Heino et al., 2023; Robinson et al., 2021). This pressure will be intensified by the expanding nutritional needs of the global human population (UN-DESA, 2024). Many have called for wide-ranging efforts to meetthese increasing food and fiber needs without further degrading the environment (Foley et al., 2011; Godfray et al., 2010; Hunter et al., 2017). Understanding how staple crops such as lentil and other pulses cope with heat and drought stress is critical to ensure global food security. Given the importance of LWMCs for stress tolerance in crops, understanding the genetic underpinnings of LMWC biosynthesis is an essential step towards developing new climate change-resilient cultivars. However, the genetic basis of LMWC biosynthesis has not been well characterized in lentil, especially related to stress tolerance. A better understanding of the genetic basis of LMWC biosynthesis will enable breeders to make more targeted selections to hasten the release of stress-tolerant lentil cultivars. The objectives of this review are to i) describe the role of LMWCs in abiotic stress tolerance, ii) summarize current knowledge of the genes involved in LMWC biosynthesis, especially in response to abiotic stress, and iii) demonstrate how LMWCs can be targeted using diverse genomic resources and markers to accelerate lentil breeding efforts for improved stress tolerance.
2 Carbohydrates in plants
Plant carbohydrates are generally classified by the type, number, and linkage configuration of monosaccharides bonded to form more complex carbohydrates (Table 2). The monosaccharides glucose, fructose, and galactose are bonded in various configurations to form polymers of increasing complexity or are reduced to form SAs. Sucrose is a disaccharide consisting of one glucose molecule and one fructose molecule; it is the primary carbon transport molecule in plants, is used to synthesize many essential compounds, and helps to mitigate abiotic stress (Huber and Huber, 1996; Kühn et al., 1999; Nägele et al., 2012; Peshev et al., 2013). Other disaccharides such as trehalose and maltose are common in plants; trehalose metabolism is tightly linked with sucrose metabolism (Lunn et al., 2014), and maltose is a starch breakdown product that is important in many aspects of carbon metabolism (Fincher, 1989; Lu and Sharkey, 2006). Oligosaccharides are carbohydrates composed of three to 20 polymerized monosaccharides (Cummings and Stephen, 2007) and have diverse physiological roles in plants, such as carbon storage, stress tolerance, and carbon transport in certain taxa (Hannah et al., 2006; Yan et al., 2022). Among oligosaccharides, RFOs and FOS are the two most abundant classes in plants (Van den Ende, 2013). Sugar alcohols such as sorbitol and mannitol are derived from glucose or fructose by one or more chemical reduction steps and have many functions in plants, including carbon transport and storage and stress tolerance (Dumschott et al., 2017; Loescher and Everard, 2000). Starch is referred to as a high molecular weight carbohydrate and is comprised of two highly polymerized carbohydrates: amylopectin (70 – 85% of starch by weight; degree of polymerization ca. 40 – 50) and amylose (15 – 30% of starch; degree of polymerization ca. 30) (Cummings and Stephen, 2007). Starch is the primary form of carbon storage in plants (MacNeill et al., 2017). Resistant starch is a nutritional term referring to starch that is not readily digested because it is i) bound within a food matrix and physically inaccessible to enzyme activity, ii) inaccessible to enzyme activity due to granule type, especially when raw, iii) recrystallized after cooking and cooling (retrograded), iv) structurally modified, or v) complexed with a lipid (Cummings and Stephen, 2007; Dhital et al., 2016; Gutiérrez and Tovar, 2021).
Table 2. Classification of common carbohydrates in plants.
The biosynthetic pathways of sucrose, oligosaccharides, and SAs have been elucidated in many plant species. These begin with fructose, glucose, or galactose, which are combined or modified to form a variety of more complex or reduced carbohydrates (Figure 1) (Dumschott et al., 2017; Johnson et al., 2020; Sanyal et al., 2023; Singh et al., 2017). Sucrose is synthesized from modified forms of glucose and fructose, i.e., uridine diphosphate glucose and fructose-6-phosphate, in a reaction catalyzed by sucrose phosphate synthase to form sucrose-6-phosphate, which is then converted to sucrose by sucrose phosphatase (Winter and Huber, 2000). The biosynthesis of RFOs typically begins with the formation of galactinol from myo-inositol and uridine diphosphate-galactose, the galactose donor, and is catalyzed by galactinol synthase (GolS or GS) (Peterbauer and Richter, 2001). Raffinose is formed from sucrose, and the galactosyl is transferred from galactinol, catalyzed by raffinose synthase (RafS or RS). Stachyose is formed from raffinose and galactinol (galactosyl donor), and is catalyzed by stachyose synthase (StaS or STS). Verbascose is formed from stachyose and galactinol, and may be catalyzed by verbascose synthase or stachyose synthase (Elango et al., 2022; Lahuta et al., 2010). Biosynthesis of FOS begins with the formation of kestose from sucrose and fructose, and is catalyzed by sucrose:sucrose 1-fructosyl-transferase. Nystose is formed from kestose and fructose, catalyzed by fructan:fructan 1-fructosyltransferase (Singh et al., 2017; Vijn and Smeekens, 1999). SA biosynthesis begins, generally, with either fructose or glucose. Mannitol is formed from fructose-6-phosphate, with two intermediate forms (mannose-6-phosphate and mannitol-1-phosphate), catalyzed by mannose-6-phosphate isomerase, mannose-6-phosphate reductase, and mannitol-1-phosphate phosphatase. Sorbitol is formed from glucose-6-phosphate, with sorbitol-6-phosphate as an intermediate step and is catalyzed by aldose-6-phosphate reductase and sorbitol-6-phosphate phosphatase (Dumschott et al., 2017; Loescher and Everard, 2000). Biosynthetic pathways for LMWCs have not yet been elucidated in lentil because these pathways are common among most flowering plants (Benkeblia, 2022; Loescher and Everard, 2000; Salerno and Curatti, 2003; Sengupta et al., 2015).
Figure 1. Biosynthetic pathways of key LMWCs involved in stress tolerance: RFOs, FOS, and SAs. Myo-inositol is a SA, although its critical role in stress tolerance appears to be related to galactinol formation. Bold text indicates LMWCs directly involved in stress response, their immediate precursors, and their key enzymes. Abbreviations: DP, degree of polymerization; DHAP, dihydroxyacetone phosphate; UDP, uridine diphosphate; Fru, fructose; Glu, glucose; Suc, sucrose; P, phosphate; PGM, phosphoglucomutase; UGPase, UDP-glucose-pyrophosphorylase; SPS, sucrose phosphate synthase; SPP, sucrose phosphatase; GolS, galactinol synthase; RafS, raffinose synthase; StaS, stachyose synthase; VerS, verbascose synthase; 1-SST, sucrose:sucrose 1-fructosyl-transferase; 1-FFT, fructan:fructan 1-fructosyltransferase; M6PI, mannose-6-phosphate isomerase; M6PR, mannose-6-phosphate reductase; M1PP, mannitol-1-phosphate phosphatase; A6PR, aldose-6-phosphate reductase; S6PP, sorbitol-6-phosphate phosphatase; MIPS, myoinositol-1-phosphate synthase; IMP, inositol mono phosphatase; GALE, UDP-galactose 4-epimerase and UDP-glucose 4-epimerase. Figure created from Dumschott et al. (2017); Johnson et al. (2020); Sanyal et al. (2023), and Singh et al. (2017).
3 Low molecular weight carbohydrates and abiotic stress
Among LMWCs, oligosaccharides and SAs are emerging as key compounds that plants use to manage abiotic stresses, such as extreme temperatures, drought, salinity, and oxidative damage (Cacela and Hincha, 2006; Corbineau et al., 2000; Shimosaka and Ozawa, 2015; Stoyanova et al., 2011) (Figure 2). The main oligosaccharides produced by lentil are RFOs, with only small amounts of FOS produced by these crops (Johnson et al., 2015, 2021) (Table 1).
Figure 2. Conceptual diagram of the diverse roles of LMWCs in abiotic stress tolerance in plants.
3.1 Osmo-protection
Water deficit stress can occur when plants are subjected to high heat, drought, or saline conditions, negatively affecting plant growth and ultimately leading to reduced crop yield and nutritional content (Barnabás et al., 2008; Choukri et al., 2020; Sehgal et al., 2018). Plants have evolved complex physiological and biochemical mechanisms to ameliorate water deficit stress, including stomatal closure, lower photosynthesis rates, and accumulation of small organic molecules in cells that maintain membrane integrity and osmotic pressure, among many others (Gururani et al., 2015; Hincha et al., 2003; Liang et al., 2020).
The potential osmo-protective role of disaccharides and oligosaccharides has been studied in vitro using a model membrane system (Crowe et al., 1984; Hincha et al., 2003). These studies demonstrate that sucrose, trehalose, and several oligosaccharides can reduce membrane desiccation damage by two potential mechanisms (Hincha et al., 2003; Koster, 1991). First, the water replacement hypothesis proposes that hydroxyl groups on LMWCs form hydrogen bonds with lipid headgroups in membranes, stabilizing membranes during dehydration and minimizing leakage (Hincha et al., 2003). Second, the cytoplasmic vitrification hypothesis proposes that the accumulation of LMWCs in cells leads to the formation of “sugar glass,” which immobilizes membranes and cytoplasmic macromolecules, protecting them from damage (Cacela and Hincha, 2006; Koster, 1991).
LMWC accumulation in response to water deficit has been observed in many plant species, including lentil. Disaccharide and oligosaccharide accumulation has been observed during drought conditions in the seeds of maize (Zea mays; Koster and Leopold, 1988; Mohammadkhani and Heidari, 2008), soybean (Glycine max; Blackman et al., 1992), field pea (Pisum sativum; Corbineau et al., 2000), and beech (Fagus sylvatica; Pukacka et al., 2009), and is thought to confer desiccation tolerance to seeds (Corbineau et al., 2000; Koster and Leopold, 1988). Drought conditions have also led to disaccharide or oligosaccharide accumulation in the stems or leaves of lentil (Foti et al., 2021), wheat (Triticum aestivum; Hou et al., 2018; Zhang et al., 2015) and chickpea (Cicer arietinum; Salvi et al., 2018), as well as the roots of chicory (Cichorium intybus; De Roover et al., 2000) and Vernonia herbacea (Garcia et al., 2011), suggesting a critical role of disaccharides and oligosaccharides in managing drought stress in vegetative tissues. The accumulation of SAs in vegetative tissues in response to drought or salt stress has also been observed in the leaves of soybean (Streeter et al., 2001), rice bean (Vigna umbellata; Wanek et al., 1997), chickpea (Orthen et al., 2000), and kiwi (Actinidia deliciosa; Klages et al., 1999). In lentil, several studies have linked drought and/or heat stress with higher concentrations of LMWCs. Foti et al. (2021) found that drought stress increased the concentrations of the disaccharide α,α-trehalose and D-myo-inositol phosphate (an RFO precursor) in a drought-tolerant genotype. Other studies have found that total LMWC or RFO concentrations in lentil seeds were linked to high temperatures and/or low precipitation, possibly in response to heat or drought stress (Johnson et al., 2015a; Graham et al., 2017). These insights point to the need for targeted research into the potential osmo-protective role of LMWCs in lentil to inform future breeding efforts.
3.2 Antioxidants
Another role of LMWCs in abiotic stress tolerance is reducing oxidative damage. Reactive oxygen species (ROS) are byproducts of plant metabolism that can damage proteins, lipids, and nucleic acids at high concentrations. These damaging compounds primarily consist of hydroxyl radicals (OH•), superoxide ion radicals (O2• –), singlet oxygen (1O2), and hydrogen peroxide (H2O2; Peshev and Van Den Ende, 2013). Plants employ various antioxidant mechanisms to scavenge ROS: vitamins C and E, enzyme-based systems such as catalase superoxidase dismutase (among others), and several secondary metabolites such as carotenoids, flavonoids, and terpenoids (Gill and Tuteja, 2010). Under optimal conditions, ROS are scavenged at the same rate they are produced by plant metabolic processes. During stress, however, the ability of plants to use these antioxidant mechanisms is diminished, leading to oxidative damage. Recent research demonstrates the role of several LMWCs in scavenging ROS to limit oxidative damage (Matros et al., 2015; Van Den Ende and Valluru, 2009). Many plant-derived disaccharides (e.g., sucrose, trehalose, and maltose), RFOs, and FOS scavenge ROS with varying degrees of affinity. In general, monosaccharides such as glucose and fructose are not effective ROS scavengers compared to disaccharides, oligosaccharides, or SAs (Morelli et al., 2003; Peshev et al., 2013; Stoyanova et al., 2011). Peshev et al. (2013) showed a 10-fold difference in ROS-scavenging capacity between trehalose (lowest capacity among carbohydrates tested) and inulin (a FOS; highest capacity). In Matros et al. (2015), A. thaliana plants were supplied with sucralose, a synthetic sucrose analog, and demonstrated a carbohydrate-antioxidant mechanism that decreased oxidative stress induced by paraquat and UV-B. While this experimental work is currently limited to A. thaliana and select vegetable crops, the scavenging of ROS by LMWCs may extend to other plant species, including lentil.
4 Gene identification and function: biosynthesis of low molecular weight carbohydrates
Much research has been carried out to identify the genes responsible for the biosynthesis of LMWCs in response to abiotic stress. To achieve this, studies have inserted, eliminated, or modified genes responsible for RFO, FOS, and SA biosynthetic pathways in many plant species, with corresponding changes in LMWC concentrations and abiotic stress tolerance (Bie et al., 2012; Egert et al., 2013; Panikulangara et al., 2004; Zhifang and Loescher, 2003).
4.1 Oligosaccharides
A critical step in the biosynthesis of RFOs is the formation of galactinol from myo-inositol and UDP-galactose, which is catalyzed by the enzyme GolS. Thus, manipulating GolS genes can affect downstream RFO concentrations and abiotic stress tolerance. For example, Panikulangara et al. (2004) found that upregulating GolS genes in A. thaliana increased raffinose concentrations in leaves and improved tolerance to heat and drought stress. In contrast, eliminating GolS genes had the opposite effect on raffinose and stress tolerance. Another study overexpressed GolS genes from chickpea (CaGolS1 and CaGolS2) in A. thaliana, which led to higher RFO concentrations in vegetative tissues and fewer signs of stress in the face of elevated temperatures (Salvi et al., 2018). Nishizawa et al. (2008) used a different approach to increase RFOs via GolS, where they overexpressed heat shock transcription factor A2 (HsfA2) to induce the transcription of GolS in A. thaliana. This led to increased raffinose concentrations in leaves and reduced oxidative damage. Putative GolS genes have also been identified in lentil (LcGolS1 and LcGolS2) using a cDNA library prepared from developing seeds, where nucleotide sequences were aligned from Medicago sativa, field pea, soybean, and Ammopiptanthus mongolicus (Kannan et al., 2016). However, follow-up work is needed to confirm these genes and their role in RFO biosynthesis in response to abiotic stress. For example, a transformation system could be employed to overexpress and suppress putative LcGolS genes, followed by quantification of gene expression, galactinol and RFOs to confirm gene identity. Similarly, the role of RFOs in abiotic stress tolerance in lentil could be further elucidated by exposing transformed plants to abiotic stress, and any differences in RFO concentrations and crop performance between transformed genotypes (overexpressing vs. suppressing LcGolS) would provide information about gene function in response to abiotic stress.
The next step in the biosynthetic pathway of RFOs is raffinose synthesis, catalyzed by RafS. Functional studies of RafS genes in many plant species have confirmed the important role of RFOs in abiotic stress tolerance (Li et al., 2020). Egert et al. (2013) demonstrated in A. thaliana using two loss-of-function mutants that a raffinose synthase gene (AtRafS5) is solely responsible for raffinose accumulation in seeds and leaves in response to drought, salinity, and oxidative stress. Li et al. (2020) demonstrated a raffinose synthase gene from maize (ZmRafS) is induced by drought, heat and salinity stress, and that a maize mutant lacking ZmRafS is drought sensitive compared to a maize null-segregant with this gene. They also found overexpression of ZmRafS in A. thaliana resulted in enhanced drought tolerance and increased raffinose concentrations in seeds. Similar to putative GolS genes in lentil (Kannan et al., 2016), putative genes for RafS and StaS identified using cDNA library have also been identified in lentil (Kannan et al., 2021); significant follow-up work is needed to confirm the presence of these genes and their function when faced with abiotic stress.
Fructooligosaccharides also have a role in stress tolerance. Transgenic studies show exotic genes responsible for FOS biosynthesis from bacteria (Bacillus subtilis) and several plant species (wheat, Jerusalem artichoke (Helianthus tuberosus), onion (Allium cepa), and Psathyrostachys huashanica) result in increased FOS concentrations and improved drought tolerance in tobacco (Bie et al., 2012; He et al., 2015; Pilon-Smits et al., 1995; Sun et al., 2020), sugar beet (Pilon-Smits et al., 1999), and cotton (Liu et al., 2022). Fructooligosaccharide concentrations in lentil seeds are considerably lower than RFO concentrations (ca. 1 – 3% of total oligosaccharides; Johnson et al., 2015), and therefore are likely less important for stress tolerance compared to RFOs.
These studies have identified the genes responsible for oligosaccharide biosynthesis in many plant species and suggest GolS and RafS genes are good candidates for increasing RFOs in lentil in order to improve tolerance to abiotic stress (Egert et al., 2013; Ma et al., 2021; Salvi et al., 2018). Given that putative genes for GolS and RafS have been identified in lentil, with better-described GolS and RafS genes in pea and chickpea (Lahuta et al., 2014; Salvi et al., 2018), further research is clearly needed to confirm the identity and function of GolS and RafS genes in lentil to support breeding efforts targeting abiotic stress tolerance. Genes encoding enzymes involved in the biosynthesis of RFOs in lentil and other plant species are shown in Table 3.
Table 3. Genes encoding enzymes involved in the biosynthesis of LMWCs in lentil and other plants.
4.2 Sugar alcohols
The role of SAs in abiotic stress mitigation has not been studied as well as that of other LMWCs, but several transgenic studies demonstrate the important role of SAs in stress tolerance. For example, genes responsible for mannitol biosynthesis (mannitol-1-phosphate dehydrogenase) transgenically introduced from E. coli improved drought and salinity stress in basmati rice (Oryza sativa; Pujni et al., 2007), and mannose-6-phosphate reductase genes introduced from celery (Apium graveolens) improved tolerance to salinity stress in A. thaliana (Zhifang and Loescher, 2003). Transgenes responsible for D-ononitol biosynthesis in a salt-tolerant rice were introduced to tobacco and increased both D-ononitol and tolerance to abiotic stresses (drought and salinity) (Sheveleva et al., 1997). Similarly, genes responsible for myo-inositol biosynthesis were introduced to tobacco, leading to increased tolerance to salinity stress (Majee et al., 2004).
Genes responsible for SA biosynthesis in lentil have not been characterized, and further research is required to understand the potential genetic underpinnings of SA biosynthesis in response to stress. Because the major SAs in lentil seeds are sorbitol and mannitol (ca. 86% and 13%, respectively; Johnson et al., 2013, 2015, 2021), research should focus on enzymes within these presumed biosynthetic pathways in lentil (Figure 1). Further, aldose-6-phosphate reductase (A6PR) and sorbitol-6-phosphate phosphatase (S6PP) may be ideal starting points, given sorbitol is considerably higher than mannitol in lentil (Table 1). Genes that encode for enzymes involved in sorbitol biosynthesis in plants are understudied, with the exception of Rosaceous tree fruits, and need further clarification in lentil and other annual crop species (Table 3).
5 Breeding potential, targets, and future directions
LMWCs in lentil and other pulse crops have been studied for crop improvement (Johnson et al., 2021; Thavarajah et al., 2022). Within the last decade, several studies have confirmed the genetic basis for variation of RFOs and SAs, suggesting these LMWCs in lentil can be increased with targeted breeding (Johnson et al., 2015, 2021). Broad-sense heritability estimates (H2) for LMWCs in lentil have not been well defined, but Johnson et al. (2021) found values ranged from 0.29 – 0.41 for RFOs and 0.34 – 0.45 for SAs within a diverse population of 143 lentil accessions (Table 1). These values are similar to those reported for other pulse crops: values for RFOs ranged from 0.25 – 0.56 in chickpea (Gangola et al., 2013) and 0.44 – 0.54 in common bean (McPhee et al., 2002). Similarly, Thavarajah et al. (2022) studied the heritability of LMWCs in field peas and found H2 values ranging from 0.64 – 0.74 for RFOs and 0.42 – 0.66 for SAs. Such moderate heritability values for LMWCs in lentil and related crops are likely due to the quantitative nature of these diverse traits. These results suggest conventional breeding efforts should be paired with genomic approaches to improve selection accuracy and efficiency to accelerate the development of new stress-tolerant lentil cultivars.
Modern genomic-assisted breeding has improved the quality and quantity of genetic data available to plant breeders to develop more climate change-resilient cultivars. This is a powerful approach because it allows the selection of parents based on higher-resolution genomic data and sophisticated statistical techniques that identify genomic regions associated with desired traits. For example, genome-wide association studies (GWAS) can aid in the identification of genes by first identifying single nucleotide polymorphisms (SNPs) and quantitative trait loci (QTL) associated with specific traits. Confirmed genes, SNPs, and QTL can then be used to improve the accuracy of selecting breeding parents by avoiding selection based solely on phenotypic information. Genomic-assisted breeding is especially useful for complex, quantitative traits that are influenced by environmental factors (Kumar et al., 2016). Thus, breeding programs that utilize genomic-assisted techniques such as GWAS can make more targeted crosses and increase genetic gain more quickly than conventional breeding programs (Figure 3).
Figure 3. Simplified genomic-assisted breeding schematic, where diverse genomic resources are phenotyped across diverse environments, followed by association studies to correlate traits with genomic regions, which can inform parent selection for crosses, leading to improved germplasm.
While the genomic regions responsible for the biosynthesis of LWMCs in lentil have received limited study, important progress toward gene identification has been made (Johnson et al., 2021; Kannan et al., 2016, 2021). Specifically, Kannan et al. (2016, 2021) identified putative GolS, RafS, and StaS genes from a cDNA library, and SNPs for mannitol and the sum of raffinose and stachyose have been identified using genome-wide association mapping (Johnson et al., 2021). These findings hold promise for continued elucidation of the genetic basis of LMWC biosynthesis in lentil, especially as more diverse genomic resources are characterized. As genomic resources for lentil are built, increased diversity will improve the predictive ability of statistical techniques used to identify candidate genes, and the diversity of potential parents will improve as well. Once QTL or genes involved in LMWC biosynthesis are better understood in lentil, more research will be required to confirm their function. For example, up- or downregulating GolS and RafS genes in lentil will help to confirm gene function, as determined by differences in RFO concentrations. Further, abiotic stress studies should be conducted in coordination with gene studies to improve our understanding of gene function and the concomitant role of LMWCs in stress tolerance in lentil. Specifically, testing crop performance as abiotic stresses are applied to lentil genotypes with up- or downregulating GolS and RafS genes will help to elucidate gene identify and function.
Within the context of a genomic-assisted breeding program, target LMWC concentrations should be developed. Target LMWC concentrations in lentil should consider their potential benefits for both human and plant health, as well as the potential drawbacks of consuming large amounts of oligosaccharides and SAs, which can lead to gastrointestinal distress in certain populations or individuals (Douglas and Sanders, 2008). While concentrations have not been established to maximize benefits for human or plant health, mean RFO (6.11 g/100 g) and SA (1.68 g/100 g) concentrations of nine commercial cultivars field-grown in six countries were below the recommended daily allowance (RDA) values of 7-30 g/day suggested for oligosaccharides (Coussement, 1999; Johnson et al., 2013; Silk et al., 2009). For sensitive individuals, these concentrations may lead to gastrointestinal discomfort but are at the low end of suggested RDA values and will likely not negatively affect most individuals.
Concentrations for LMWCs such as RFOs and SAs in vegetative tissue, which are important to consider given their critical role throughout the crop’s life cycle, are not often measured because most research has focused on the nutrient content and digestibility of lentil seeds. Thus, target concentrations for LMWCs in lentil vegetative tissues will require further study to identify optimal concentrations under different environmental conditions, especially abiotic stress. Any physiological tradeoffs between LMWC biosynthesis and other aspects of carbon metabolism should also be considered when determining target LMWC concentrations.
To make efficient progress toward new stress-tolerant lentil cultivars, future work must rely on diverse genomic resources and recent advances in genomic techniques such as whole-genome sequencing and complimentary statistical analyses. Yet, more research is needed to better understand the genes responsible for LMWC biosynthesis in lentil, as well optimal quantities for both human and plant health. With this information, genomic-assisted breeding techniques can be employed to hasten the development of stress tolerant lentil cultivars for a more food secure future.
6 Conclusion
Lentil is a nutrient-dense food crop that is well adapted to the challenging growing conditions of the dry regions where it is primarily produced, and it is well positioned to contribute to global food security in the future. However, more research is needed to take better advantage of LMWCs in lentil to improve tolerance to heat and drought stress, thereby addressing the dual threats posed by climate change and a growing human population. Importantly, LMWCs in lentil also provide health benefits to consumers, which is significant for all global consumers. Broadening and characterizing lentil genomic resources are the first steps toward better understanding the genes responsible for LMWC biosynthesis in lentil, followed by confirming gene function as stress response compounds. From this work, diverse genomic resources and genomic-assisted breeding techniques can be leveraged to develop more climate-resilient lentil cultivars based on LMWCs. Thus, by employing genomic-assisted breeding techniques that focus on LMWCs, lentil yield and nutritional quality can be maintained or improved, helping to ensure food security in a changing world.
Author contributions
MD: Conceptualization, Data curation, Formal analysis, Methodology, Writing – original draft, Writing – review & editing. DT: Conceptualization, Data curation, Funding acquisition, Investigation, Project administration, Resources, Supervision, Writing – review & editing.
Funding
The author(s) declare financial support was received for the research, authorship, and/or publication of this article. Funding support for this project was provided by the Organic Agriculture Research and Extension Initiative (OREI); the United States Department of Agriculture; the National Institute of Food and Agriculture (award no./proposal no. 2021-02927) (DT); the USDA National Institute of Food and Agriculture [Hatch] project [1022664] (DT); USDA-ARS (DT); SC Department of Agriculture; and FoodShot Global.
Conflict of interest
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.
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Keywords: pulse crops, biofortification, lentil, low molecular weight carbohydrates, raffinose family oligosaccharides, sugar alcohols, abiotic stress
Citation: Dempsey M and Thavarajah D (2024) Low molecular weight carbohydrates and abiotic stress tolerance in lentil (Lens culinaris Medikus): a review. Front. Plant Sci. 15:1408252. doi: 10.3389/fpls.2024.1408252
Received: 27 March 2024; Accepted: 13 September 2024;
Published: 03 October 2024.
Edited by:
Ilaria Marcotuli, University of Bari Aldo Moro, ItalyReviewed by:
Aamir Raina, Aligarh Muslim University, IndiaUday Chand Jha, Indian Institute of Pulses Research (ICAR), India
Javaid Akhter Bhat, Nanjing Agricultural University, China
Copyright © 2024 Dempsey and Thavarajah. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Dil Thavarajah, ZHRoYXZhckBjbGVtc29uLmVkdQ==