Localised expression of OsIAA29 suggests a key role for auxin in regulating development of the dorsal aleurone of early rice grains

Endosperm of rice and other cereals accumulates high concentrations of the predominant in planta auxin, indole-3-acetic acid (IAA) during early grain development. However, IAA signalling and function during endosperm development are poorly understood. Here, we report that OsYUC12 (an auxin biosynthesis gene) and OsIAA29 (encoding a non-canonical AUX/IAA) are both expressed exclusively in grains, reaching a maximum 5 to 6 days after pollination. OsYUC12 expression is localized in the aleurone, sub-aleurone and embryo, whereas OsIAA29 expression is restricted to a narrow strip in the dorsal aleurone, directly under the vascular bundle. Although rice has been reported to lack endosperm transfer cells (ETCs), this region of the aleurone is enriched with sugar transporters and is likely to play a key role in apoplastic nutrient transfer, analogous to ETCs in other cereals. OsIAA29 has orthologues only in grass species; expression of which is also specific to early grain development. OsYUC12 and OsIAA29 are temporally co-expressed with two genes (AL1 and OsPR602) previously linked to the development of dorsal aleurone or ETCs. Also up regulated at the same time are a cluster of MYB-related genes (designated OsMRPLs) homologous to ZmMRP-1, which regulates maize ETC development. Wheat homologues of ZmMRP-1 are also expressed in ETCs. Although previous work has suggested that other cereals do not have orthologues of ZmMRP-1, our work suggests OsIAA29 and OsMRPLs and their homologues in other grasses are part of an auxin-regulated, conserved signalling network involved in the differentiation of cells with ETC-like function in developing cereal grains.Main ConclusionNon-canonical AUX/IAA protein, OsIAA29, and ZmMPR-1 homologues, OsMRPLs, are part of an auxin-related signalling cascade operating in the dorsal aleurone during early rice grain development.

Wheat amino acid transporters highly expressed in grain cells regulate amino acid accumulation in grain

Amino acids are delivered into developing wheat grains to support the accumulation of storage proteins in the starchy endosperm, and transporters play important roles in regulating this process. RNA-seq, RT-qPCR, and promoter-GUS assays showed that three amino acid transporters are differentially expressed in the endosperm transfer cells (TaAAP2), starchy endosperm cells (TaAAP13), and aleurone cells and embryo of the developing grain (TaAAP21), respectively. Yeast complementation revealed that all three transporters can transport a broad spectrum of amino acids. RNAi-mediated suppression of TaAAP13 expression in the starchy endosperm did not reduce the total nitrogen content of the whole grain, but significantly altered the composition and distribution of metabolites in the starchy endosperm, with increasing concentrations of some amino acids (notably glutamine and glycine) from the outer to inner starchy endosperm cells compared with wild type. Overexpression of TaAAP13 under the endosperm-specific HMW-GS (high molecular weight glutenin subunit) promoter significantly increased grain size, grain nitrogen concentration, and thousand grain weight, indicating that the sink strength for nitrogen transport was increased by manipulation of amino acid transporters. However, the total grain number was reduced, suggesting that source nitrogen remobilized from leaves is a limiting factor for productivity. Therefore, simultaneously increasing loading of amino acids into the phloem and delivery to the spike would be required to increase protein content while maintaining grain yield.

Localization, Gene Expression, and Functions of Glutamine Synthetase Isozymes in Wheat Grain (Triticum aestivum L.)

Glutamine synthetase (GS) plays a major role in plant nitrogen metabolism, but the roles of individual GS isoforms in grains are unknown. Here, the localization and expression of individual TaGS isozymes in wheat grain were probed with TaGS isoenzyme-specific antibodies, and the nitrogen metabolism of grain during the grain filling stage were investigated. Immunofluorescence revealed that TaGS1;1, TaGS1;3, and TaGS2 were expressed in different regions of the embryo. In grain transporting tissues, TaGS1;2 was localized in vascular bundle; TaGS1;2 and TaGS1;1 were in chalaza and placentochalaza; TaGS1;1 and TaGS1;3 were in endosperm transfer cells; and TaGS1;3 and TaGS2 were in aleurone layer. GS exhibited maximum activity and expression at 8 days after flowering (DAF) with peak glutamine content in grains; from then, NH4+ increased largely from NO3- reduction, glutamate dehydrogenase (GDH) aminating activity increased continuously, and the activities of GS and glutamate synthase (GOGAT) decreased, while only TaGS1;3 kept a stable expression in different TaGS isozymes. Hence, GS-GOGAT cycle and GDH play different roles in NH4+ assimilation of grain in different stages of grain development; TaGS1;3, located in aleurone layer and endosperm transfer cells, plays a key role in Gln into endosperm for gluten synthesis. At 30 DAF, grain amino acids are mainly transported from maternal phloem.