Fig. 1. Role of amino acid transporter OsAAP18 in rice yield and grain quality: insights from its haplotype variation, spatiotemporal expression, and function. A, Distribution of single nucleotide polymorphism (SNP) sites in OsAAP18 of 475 rice germplasm resources. B, Relative expression level of OsAAP18 in three haplotypes (Hap1, Hap2, and Hap3). Data are mean ± SD (n = 10). C‒E, Comparison of agronomic traits among different haplotypes: tiller number per plant (C), grain yield per plant (D), and grain width (E). Data are mean ± SD (n ≥ 10). F, Relative expression level of OsAAP18 in various tissues of Zhonghua (ZH11) during the vegetative and reproductive growth stages. Data are mean ± SD (n = 3). Transcript levels were quantified by qPCR using the rice OsActin1 gene as an internal control. G and H, Cross sections of leaf sheath (G) and young panicle (H) from OsAAP18 promoter-β-glucuronidase gene (GUS) transgenic plants. VT, Vascular tissue. Scale bars, 50 μm. I, Subcellular localization and co-localization of OsAAP18 with mCherry-fused cell membrane protein OsMCA1. GFP, Green fluorescent protein. Scale bars, 10 μm. J, Growth of yeast strains cultured at 30 ºC for 48 h on solid medium containing 1 mmol/L (NH4)2SO4 or 1 mmol/L asparagine (Asn), glycine (Gly), leucine (Leu), isoleucine (Ile), proline (Pro), phenylalanine (Phe), threonine (Thr), or valine (Val) as the sole nitrogen source. The wild-type strain 23344c was used as the positive control, and the yeast mutant 22Δ10α expressing the empty vector pDR196 was used as the negative control. K and L, Phenotypes of mature plants (K) and grain yield per plant (L) of ZH11, OsAAP18-OE (overexpression) lines (OE1‒3), and OsAAP18-C (CRISPR/Cas9 mutant) lines (C1‒3). Scale bars are 15 cm in K and 2 cm in L. M‒Q, Yield-related traits: tiller number per plant (M), grain yield per plant (N), filled grain number per plant (O), 1000-grain weight (P), and seed-setting rate (Q) of ZH11, OsAAP18-OE, and OsAAP18-C lines. Data are mean ± SD (n = 20). R, Contents of free amino acids in mature grains of ZH11, OsAAP18-OE, and OsAAP18-C lines. Data are mean ± SD (n = 3). Asp, Aspartic acid; Ser, Serine; Glu, Glutamic acid; Ala, Alanine; Cys, Cysteine; Met, Methionine; Tyr, Tyrosine; His, Histidine; Lys, Lysine; Arg, Arginine. S, Grain widths of brown rice for ZH11, OsAAP18-OE, and OsAAP18-C lines. Scale bars, 5 mm. Data are mean ± SD (n ≥ 500). T‒V, Grain quality parameters: protein content (T), total starch content (U), and amylose content (V) of ZH11, OsAAP18-OE, and OsAAP18-C lines. Data are mean ± SD (n = 3). Different lowercase letters above bars indicate significant differences at the P < 0.05 level by Duncan’s multiple comparison test. *, **, and *** indicate significant differences at P < 0.05, P < 0.01, and P < 0.001, respectively. ns, Not significant.
Fig. 1. Role of amino acid transporter OsAAP18 in rice yield and grain quality: insights from its haplotype variation, spatiotemporal expression, and function. A, Distribution of single nucleotide polymorphism (SNP) sites in OsAAP18 of 475 rice germplasm resources. B, Relative expression level of OsAAP18 in three haplotypes (Hap1, Hap2, and Hap3). Data are mean ± SD (n = 10). C‒E, Comparison of agronomic traits among different haplotypes: tiller number per plant (C), grain yield per plant (D), and grain width (E). Data are mean ± SD (n ≥ 10). F, Relative expression level of OsAAP18 in various tissues of Zhonghua (ZH11) during the vegetative and reproductive growth stages. Data are mean ± SD (n = 3). Transcript levels were quantified by qPCR using the rice OsActin1 gene as an internal control. G and H, Cross sections of leaf sheath (G) and young panicle (H) from OsAAP18 promoter-β-glucuronidase gene (GUS) transgenic plants. VT, Vascular tissue. Scale bars, 50 μm. I, Subcellular localization and co-localization of OsAAP18 with mCherry-fused cell membrane protein OsMCA1. GFP, Green fluorescent protein. Scale bars, 10 μm. J, Growth of yeast strains cultured at 30 ºC for 48 h on solid medium containing 1 mmol/L (NH4)2SO4 or 1 mmol/L asparagine (Asn), glycine (Gly), leucine (Leu), isoleucine (Ile), proline (Pro), phenylalanine (Phe), threonine (Thr), or valine (Val) as the sole nitrogen source. The wild-type strain 23344c was used as the positive control, and the yeast mutant 22Δ10α expressing the empty vector pDR196 was used as the negative control. K and L, Phenotypes of mature plants (K) and grain yield per plant (L) of ZH11, OsAAP18-OE (overexpression) lines (OE1‒3), and OsAAP18-C (CRISPR/Cas9 mutant) lines (C1‒3). Scale bars are 15 cm in K and 2 cm in L. M‒Q, Yield-related traits: tiller number per plant (M), grain yield per plant (N), filled grain number per plant (O), 1000-grain weight (P), and seed-setting rate (Q) of ZH11, OsAAP18-OE, and OsAAP18-C lines. Data are mean ± SD (n = 20). R, Contents of free amino acids in mature grains of ZH11, OsAAP18-OE, and OsAAP18-C lines. Data are mean ± SD (n = 3). Asp, Aspartic acid; Ser, Serine; Glu, Glutamic acid; Ala, Alanine; Cys, Cysteine; Met, Methionine; Tyr, Tyrosine; His, Histidine; Lys, Lysine; Arg, Arginine. S, Grain widths of brown rice for ZH11, OsAAP18-OE, and OsAAP18-C lines. Scale bars, 5 mm. Data are mean ± SD (n ≥ 500). T‒V, Grain quality parameters: protein content (T), total starch content (U), and amylose content (V) of ZH11, OsAAP18-OE, and OsAAP18-C lines. Data are mean ± SD (n = 3). Different lowercase letters above bars indicate significant differences at the P < 0.05 level by Duncan’s multiple comparison test. *, **, and *** indicate significant differences at P < 0.05, P < 0.01, and P < 0.001, respectively. ns, Not significant.
