In plants, myo-inositol-1,2,3,4,5,6-hexakisphosphate (InsP6), also known as phytic acid (PA), is a major component of organic phosphorus (P), and accounts for up to 85% of the total P in seeds. In rice (Oryza sativa L.), PA mainly accumulates in rice bran, and chelates mineral cations, resulting in mineral deficiencies among brown rice consumers. Therefore, considerable efforts have been focused on the development of low PA (LPA) rice cultivars. In this study, we performed genetic and molecular analyses of OsLpa1, a major PA biosynthesis gene, in Sanggol, a low PA mutant variety developed via chemical mutagenesis of Ilpum rice cultivar. Genetic segregation and sequencing analyses revealed that a recessive allele, lpa1-3, at the OsLpa1 locus (Os02g0819400) was responsible for a significant reduction in seed PA content in Sanggol. The lpa1-3 gene harboured a point mutation (C623T) in the fourth exon of the predicted coding region, resulting in threonine (Thr) to isoleucine (Ile) amino acidsubstitution at position 208 (Thr208Ile). Three-dimensional analysis of Lpa1 protein structure indicated that myo-inositol 3-monophosphate [Ins(3)P1] could bind to the active site of Lpa1, with ATP as a cofactor for catalysis. Furthermore, the presence of Thr208 in the loop adjacent to the entry site of the binding pocket suggests that Thr208Ile substitution is involved in regulating enzyme activity via phosphorylation. Therefore, we propose that Thr208Ile substitution in lpa1-3 reduces Lpa1 enzyme activity in Sanggol, resulting in reduced PA biosynthesis.

Background Although embryo accounts for only 2–3% of the total weight of a rice grain, it is a good source of various nutrients for human health. Because enlarged embryo size causes increase of the amount of nutrients and bioactive compounds stored within rice grain, giant embryo mutants of rice ( Oryza sativa L.) are excellent genetic resources for improving the nutritional value of rice grains. Results Three giant embryo mutants, including large embryo ( le ), giant embryo ( ge ) and super - giant embryo ( ge s ), with variable embryo size were used in this study. We investigated whether genes controlling embryo size in these mutants ( le , ge and ge s ) were allelic to each other. Although ge and ge s was allelic to GIANT EMBRY ( GE ), le was not allelic to ge and ge s in allelism test. The GE gene carried a unique nucleotide substitution in each of the two mutants ( ge and ge s ), resulting in non-synonymous mutations in exon 2 of GE in both mutants. However, the GE gene of the le mutant did not carry any mutation, suggesting that the enlarged embryo phenotype of le was governed by another gene. Using map-based cloning, we mapped the LE gene to the short arm of chromosome 3. The le mutant showed mild enlargement in embryo size, which resulted from an increase in the size of scutellar parenchyma cells. The LE encodes a C3HC4-type RING finger protein and was expressed to relatively high levels in seeds at a late developmental stage. Knockdown of LE expression using RNA interference increased the embryo size of rice grains, confirming the role of LE in determining the embryo size. Conclusion Overall, we identified a new gene controlling embryo size in rice. Phenotypic and molecular characterization results suggest that the le mutant will serve as a valuable resource for developing new rice cultivars with large embryos and nutrient-dense grains. Electronic supplementary material The online version of this article (10.1186/s12284-019-0277-y) contains supplementary material, which is available to a...

21In plants, 2,3,4,5, ), also known as phytic acid 22 (PA), is a major component of organic phosphorus (P), and accounts for up to 85% of the total 2 23 P in seeds. In rice (Oryza sativa L.), PA mainly accumulates in rice bran, and chelates mineral 24 cations, resulting in mineral deficiencies among brown rice consumers. Therefore, 25 considerable efforts have been focused on the development of low PA (LPA) rice cultivars. In 26 this study, we performed genetic and molecular analyses of OsLpa1, a major PA biosynthesis 27 gene, in Sanggol, a low PA mutant variety developed via chemical mutagenesis of Ilpum rice 28 cultivar. Genetic segregation and sequencing analyses revealed that a recessive allele, lpa1-3, 29 at the OsLpa1 locus (Os02g0819400) was responsible for a significant reduction in seed PA 30 content in Sanggol. The lpa1-3 gene harboured a point mutation (C623T) in the fourth exon of 31 the predicted coding region, resulting in threonine (Thr) to isoleucine (Ile) amino acid 32 substitution at position 208 (Thr208Ile). Three-dimensional analysis of Lpa1 protein structure 33 indicated that myo-inositol 3-monophosphate [Ins(3)P 1 ] kinase binds to the active site of Lpa1, 34 with ATP as a cofactor for catalysis. Furthermore, the presence of Thr208 in the loop adjacent 35 to the entry site of the binding pocket suggests that Thr208Ile substitution is involved in 36 regulating enzyme activity via phosphorylation. Therefore, we propose that Thr208Ile 37 substitution in lpa1-3 reduces Lpa1 enzyme activity in Sanggol, resulting in reduced PA 38 biosynthesis. 39 40 42 phytic acid (PA), is considered a major source of phosphorus (P) available in the form of 43 phytate, and accounts for 65-85% of the total P in seeds [1]. Monogastric animals poorly digest 44 PA, as they lack the phytase enzyme, which is responsible for the release of phosphate residues 45 [2]. PA is an efficient chelator of mineral cations, such as zinc (Zn 2+ ), iron (Fe 2+ ), magnesium 46 (Mg 2+ ), potassium (K 2+ ), and calcium (Ca 2+ ), in the nutritional tract. Because of these attributes, 3 47 PA is considered as an antinutrient [3, 4]. Hence, there is a need to develop low PA (LPA) crop 48 cultivars to maximize the nutritional benefits of grains. 49 Mutants associated with the LPA phenotype have been identified in several crop plants 50 including maize (Zea mays) [5, 6], barley (Hordeum vulgare) [7], soyabean (Glycine max) [8], 51 rice (Oryza sativa) [9], and wheat (Triticum aestivum) [10]. Although, LPA mutants are 52identified primarily on the basis of percentage reduction of PA and high inorganic P (P i ) content 53 in seeds [5, 11], some mutants show a significant accumulation of myo-inositol and inositol 54 phosphate [Ins(1,3,4)P 3 5-/6] intermediates in seeds [12, 13]. 55Previously, the LPA phenotype of seeds has been associated with reduced agronomic 56 performance of mutant crop plants in the field [5, 14]. It is important to understand the genetic 57 and molecular bases of reduced agronomic performance of LPA mutants for ef...

The resveratrol-producing rice (Oryza sativa L.) inbred lines, Iksan 515 (I.515) and Iksan 526 (I.526), developed by the expression of the groundnut (Arachis hypogaea) resveratrol synthase 3 (AhRS3) gene in the japonica rice cultivar Dongjin, accumulated both resveratrol and its glucoside, piceid, in seeds. Here, we investigated the effect of the AhRS3 transgene on the expression of endogenous piceid biosynthesis genes (UGTs) in the developing seeds of the resveratrol-producing rice inbred lines. Ultra-performance liquid chromatography (UPLC) analysis revealed that I.526 accumulates significantly higher resveratrol and piceid in seeds than those in I.515 seeds and, in I.526 seeds, the biosynthesis of resveratrol and piceid reached peak levels at 41 days after heading (DAH) and 20 DAH, respectively. Furthermore, RNA-seq analysis showed that the expression patterns of UGT genes differed significantly between the 20 DAH seeds of I.526 and those of Dongjin. Quantitative real-time PCR (RT-qPCR) analyses confirmed the data from RNA-seq analysis in seeds of Dongjin, I.515 and I.526, respectively, at 9 DAH, and in seeds of Dongjin and I.526, respectively, at 20 DAH. A total of 245 UGTs, classified into 31 UGT families, showed differential expression between Dongjin and I.526 seeds at 20 DAH. Of these, 43 UGTs showed more than 2-fold higher expression in I.526 seeds than in Dongjin seeds. In addition, the expression of resveratrol biosynthesis genes (PAL, C4H and 4CL) was also differentially expressed between Dongjin and I.526 developing seeds. Collectively, these data suggest that AhRS3 altered the expression pattern of UGT genes, and PAL, C4H and 4CL in developing rice seeds.