Heat Priming of Lentil (Lens culinaris Medik.) Seeds and Foliar Treatment with γ-Aminobutyric Acid (GABA), Confers Protection to Reproductive Function and Yield Traits under High-Temperature Stress Environments

Abstract

Gradually increasing temperatures at global and local scales are causing heat stress for cool and summer-season food legumes, such as lentil (Lens culinaris Medik.), which is highly susceptible to heat stress, especially during its reproductive stages of development. Hence, suitable strategies are needed to develop heat tolerance in this legume. In the present study, we tested the effectiveness of heat priming (HPr; 6 h at 35 °C) the lentil seeds and a foliar treatment of γ-aminobutyric acid (GABA; 1 mM; applied twice at different times), singly or in combination (HPr+GABA), under heat stress (32/20 °C) in two heat-tolerant (HT; IG2507, IG3263) and two heat-sensitive (HS; IG2821, IG2849) genotypes to mitigate heat stress. The three treatments significantly reduced heat injury to leaves and flowers, particularly when applied in combination, including leaf damage assessed as membrane injury, cellular oxidizing ability, leaf water status, and stomatal conductance. The combined HPr+GABA treatment significantly improved the photosynthetic function, measured as photosynthetic efficiency, chlorophyll concentration, and sucrose synthesis; and significantly reduced the oxidative damage, which was associated with a marked up-regulation in the activities of enzymatic antioxidants. The combined treatment also facilitated the synthesis of osmolytes, such as proline and glycine betaine, by upregulating the expression of their biosynthesizing enzymes (pyrroline-5-carboxylate synthase; betaine aldehyde dehydrogenase) under heat stress. The HPr+GABA treatment caused a considerable enhancement in endogenous levels of GABA in leaves, more so in the two heat-sensitive genotypes. The reproductive function, measured as germination and viability of pollen grains, receptivity of stigma, and viability of ovules, was significantly improved with combined treatment, resulting in enhanced pod number (21-23% in HT and 35-38% in HS genotypes, compared to heat stress alone) and seed yield per plant (22-24% in HT and 37-40% in HS genotypes, in comparison to heat stress alone). The combined treatment (HPr+GABA) was more effective and pronounced in heat-sensitive than heat-tolerant genotypes for all the traits tested. This study offers a potential solution for tackling and protecting heat stress injury in lentil plants.

Keywords: high temperature; legumes; lentil; pods; pollen; seeds; stress.

Figure 1

Phenology (Days to podding: (A); days to maturity: (B); flowering-podding interval: (C); podding—maturity interval: (D) of heat-tolerant (G1: IG2507; G2: IG3263) and heat-sensitive (G1: IG2821; G2: IG2849) genotypes in control (28/18 °C; 12 h each), heat-stressed (32/20 °C; 12 h each), heat-primed (HPr), GABA-treated, and HPr+GABA treatments. Plants were exposed to heat stress at the onset of flowering (bud stage) for all genotypes; hence, the phenological data related to days to flowering is similar for all treatments. Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD (least significant difference) for interaction (genotypes × stages × treatments) (p < 0.05): Days to podding: 1.8; days to maturity: 1.9; flowering-podding interval: 1.8; podding—maturity interval: 1.7.

Figure 2

Effect of heat priming (HPr) and γ-amino butyric acid (GABA), applied individually or in combination (HPr+GABA) on membrane damage (A), cellular oxidizing ability (B), relative leaf water content, RLWC (C) and stomatal conductance (D) on heat-tolerant (G1: IG2507; G2: IG3263) and heat-sensitive (G1: IG2821; G2: IG2849) genotypes at stage 1 (S1) and stage 2 (S2) in heat-stressed (HS) lentil plants, compared to control (C). Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD for interaction (genotypes × stages × treatments) (p < 0.05): membrane damage: 2.9; cellular oxidizing ability: 0.030, relative leaf water content: 2.2, stomatal conductance: 26.3. FW = fresh weight.

Figure 3

Effect of heat priming (HPr) and γ-amino butyric acid (GABA), applied individually or in combination (HPr+GABA) on pollen germination (A), pollen viability (B), stigma receptivity (C) and ovule viability (D) on heat-tolerant (G1: IG2507; G2: IG3263) and heat-sensitive (G1: IG2821; G2: IG2849) genotypes at stage 1 (S1) and stage 2 (S2) in heat-stressed (HS) lentil plants, compared to control (C). Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD for interaction (genotypes × stages × treatments) (p < 0.05): pollen germination: 6.2, pollen viability: 6.4, stigma receptivity: 0.25, ovule viability: 0.26.

Figure 4

Effect of heat priming (HPr) and γ-amino butyric acid (GABA), applied individually or in combination (HPr+GABA) on photosystem (PS) II function (A), chlorophyll content (B), sucrose (C) and sucrose phosphate synthase, SPS (D) on heat-tolerant (G1: IG2507; G2: IG3263) and heat-sensitive (G1: IG2821; G2: IG2849) genotypes at stage 1 (S1) and stage 2 (S2) in heat-stressed (HS) lentil plants, compared to control (C). Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD for interaction (genotypes × stages × treatments) (p < 0.05): photosystem II: 0.039, chlorophyll content: 1.9; sucrose: 2.3, sucrose phosphate synthase: 0.91. dw = dry weight.

Figure 5

Effect of heat priming (HPr) and γ-amino butyric acid (GABA), applied individually or in combination (HPr+GABA) on malondialdehyde, MDA (A) and hydrogen peroxide, H₂O₂ (B) on heat-tolerant (G1: IG2507; G2: IG3263) and heat-sensitive (G1: IG2821; G2: IG2849) genotypes at stage 1 (S1) and stage 2 (S2) in heat-stressed (HS) lentil plants, compared to control (C). Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD for interaction (genotypes × stages × treatments) (p < 0.05): malondialdehyde: 1.9, hydrogen peroxide: 0.74. dw = dry weight.

Figure 6

Effect of heat priming (HPr) and γ-amino butyric acid (GABA), applied individually or in combination (HPr+GABA) on superoxide dismutase (A), catalase (B), ascorbate peroxidase (C) and glutathione reductase (D) activities on heat-tolerant (G1: IG2507; G2: IG3263) and heat-sensitive (G1: IG2821; G2: IG2849) genotypes at stage 1 (S1) and stage 2 (S2) in heat-stressed (HS) lentil plants, compared to control (C). Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD for interaction (genotypes × stages × treatments) (p < 0.05): superoxide dismutase: 0.21, catalase: 0.19; ascorbate peroxidase: 0.20, glutathione reductase: 0.19.

Figure 7

Effect of heat priming (HPr) and γ-amino butyric acid (GABA), applied individually or in combination (HPr+GABA) on proline (A) and pyrolline-5- carboxylate synthase (B) glycine betaine (C) and betaine aldehyde dehydrogenase (D) on heat-tolerant (G1: IG2507; G2: IG3263) and heat-sensitive (G1: IG2821; G2: IG2849) genotypes at stage 1 (S1) and stage 2 (S2) in heat-stressed (HS) lentil plants, compared to control (C). Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD for interaction (genotypes × stages × treatments) (p < 0.05): proline: 2.7, pyrolline-5-carboxylate synthase: 2.8, glycine betaine: 2.8, betaine aldehyde dehydrogenase: 0.19. dw = dry weight.

Figure 8

Effect of heat priming (HPr) and γ-amino butyric acid (GABA), applied individually or in combination (HPr+GABA) on endogenous GABA in heat-tolerant (G1: IG2507; G2: IG3263) and heat-sensitive (G1: IG2821; G2: IG2849) genotypes at stage 1 (S1) and stage 2 (S2) in heat-stressed (HS) lentil plants, compared to control (C). Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD for interaction (genotypes × stages × treatments) (p < 0.05): 0.038. dw = dry weight.

Figure 9

Effect of heat priming (HPr) and γ-amino butyric acid (GABA), applied individually or in combination (HPr+GABA) on pod number (A) and seed yield/plant (B) in heat-tolerant (HT; G1: IG2507; G2: IG3263) and heat-sensitive (HS; G1: IG2821; G2: IG2849) genotypes in heat-stressed (HS) lentil plants, compared to control (C). Vertical bars represent standard errors (n = 3). Different small letters on the bars indicate significant differences from each other (p < 0.05). LSD for interaction (genotypes × stages × treatments) (p < 0.05): pod number: 6.8, seed yield/plant: 0.29.

Figure 10

Temperature profile (maximum (Max), minimum (Min) and average (Avg)) from sowing to onset of flowering in outdoor environment at the experimental site.