Overexpression of Rice Rab7 Gene Improves Drought and Heat Tolerance and Increases Grain Yield in Rice (Oryza sativa L.)

Abstract

Rab family proteins play a crucial role in plant developmental processes and tolerance to environmental stresses. The current study investigated whether rice Rab7 (OsRab7) overexpression could improve rice tolerance to drought and heat stress conditions. The OsRab7 gene was cloned and transformed into rice plants. The survival rate, relative water content, chlorophyll content, gas-exchange characteristics, soluble protein content, soluble sugar content, proline content, and activities of antioxidant enzymes (CAT, SOD, APX, POD) of the transgenic rice lines were significantly higher than that of the wild-type. In contrast, the levels of hydrogen peroxide, electrolyte leakage, and malondialdehyde of the transgenic lines were significantly reduced when compared to wild-type. Furthermore, the expression of four genes encoding reactive oxygen species (ROS)-scavenging enzymes (OsCATA, OsCATB, OsAPX2, OsSOD-Cu/Zn) and eight genes conferring abiotic stress tolerance (OsLEA3, OsRD29A, OsSNAC1, OsSNAC2, OsDREB2A, OsDREB2B, OsRAB16A, OsRAB16C) was significantly up-regulated in the transformed rice lines as compared to their expression in wild-type. OsRab7 overexpression also increased grain yield in rice. Taken together, the current study indicates that the OsRab7 gene improves grain yield and enhances drought and heat tolerance in transgenic rice by modulating osmolytes, antioxidants and abiotic stress-responsive genes expression. Therefore, OsRab7 gene could be exploited as a promising candidate for improving rice grain yield and stress tolerance.

Keywords: OsRab7; antioxidants; drought and heat stress; gene expression; transgenic rice.

Figure 1

Molecular analysis of the transgenic rice lines. (A) PCR amplification of 750 bp of hpt hygromycin resistant gene in the wild-type (WT) and the 5 transgenic (OE) lines. (B) Southern blot analysis of the digested genomic DNA from the WT and the 5 transgenic lines. (C) Relative expression of OsRab7 in transgenic lines using qRT-PCR analysis. (D) Survival rate of the WT and transgenic lines following 10-day recovery after drought and heat treatments. M, 100 bp DNA ladder. Different letters above the columns indicate significant differences between lines (p ≤ 0.05).

Figure 2

Hydrogen peroxide (H₂O₂, µmol g⁻¹ FW) content (A), lipid peroxidation (malondialdehyde (MDA), µmol g⁻¹ FW) level (B), and electrolyte leakage (C) of the wild-type and transgenic rice plants overexpressing OsRab7 subjected to normal, drought and heat stress conditions. Data are means ± SE (n = 3). Different letters above the columns indicate significant differences between rice lines (p ≤ 0.05). SE: Standard error.

Figure 3

Photosynthesis rate (P_n, μmol m²s⁻¹) (A), stomatal conductance (g_s, mol m²s⁻¹) (B), and transpiration rate (E, mmol m²s⁻¹) (C) of the wild-type and transgenic rice plants overexpressing OsRab7 subjected to normal, drought, and heat stress conditions. Data are means ± SE (n = 3). Different letters above the columns indicate significant differences (p ≤ 0.05).

Figure 4

Contents of total soluble protein (mg g⁻¹ FW) (A), soluble sugars (mg g⁻¹ FW) (B), proline (µg g⁻¹ FW) (C) and chlorophyll (mg g⁻¹ FW) (D) of the wild-type and transgenic rice plants overexpressing OsRab7 subjected to normal, drought and heat stress conditions. Data are means ± SE (n = 3). Different letters above the columns indicate significant differences between lines (p ≤ 0.05).

Figure 5

Activities of catalase (CAT) (A), superoxide dismutase (SOD) (B), ascorbate peroxidase (APX) (C), and peroxidase (POD) (D) in the wild-type and transgenic rice plants overexpressing OsRab7 subjected to normal, drought, and heat stress conditions. Data are means ± SE (n = 3). Different letters above the columns indicate significant differences between rice lines (p ≤ 0.05).

Figure 6

Expression levels of OsCATA (A), OsCATB (B), OsAPX2 (C), and OsSOD-Cu/Zn (D) genes in the wild-type and transgenic rice plants overexpressing OsRab7 subjected to normal, drought, and heat stress conditions. Data are means ± SE (n = 3). Different letters above the columns indicate significant differences between rice lines (p ≤ 0.05).

Figure 7

Expression levels of OsLEA3 (A), OsRD29A (B), OsSNAC1 (C), OsSNAC2 (D), OsDREB2A (E), OsDREB2B (F), OsRAB16A (G), and OsRAB16C (H) genes in the wild-type and transgenic rice plants overexpressing OsRab7 under normal, drought, and heat stress conditions. Data are means ± SE (n = 3). Different letters above columns indicate significant difference between rice lines (p ≤ 0.05).