As an essential macroelement for all living cells, phosphorus is indispensable in agricultural production systems. Natural phosphorus reserves are limited, and it is therefore important to develop phosphorus-efficient crops. A major quantitative trait locus for phosphorus-deficiency tolerance, Pup1, was identified in the traditional aus-type rice variety Kasalath about a decade ago. However, its functional mechanism remained elusive until the locus was sequenced, showing the presence of a Pup1-specific protein kinase gene, which we have named phosphorus-starvation tolerance 1 (PSTOL1). This gene is absent from the rice reference genome and other phosphorus-starvation-intolerant modern varieties. Here we show that overexpression of PSTOL1 in such varieties significantly enhances grain yield in phosphorus-deficient soil. Further analyses show that PSTOL1 acts as an enhancer of early root growth, thereby enabling plants to acquire more phosphorus and other nutrients. The absence of PSTOL1 and other genes-for example, the submergence-tolerance gene SUB1A-from modern rice varieties underlines the importance of conserving and exploring traditional germplasm. Introgression of this quantitative trait locus into locally adapted rice varieties in Asia and Africa is expected to considerably enhance productivity under low phosphorus conditions.

anthomonas oryzae pv. oryzae (Xoo) is the etiological agent of bacterial blight disease in rice. The disease is most severe in southeast Asia but is increasingly damaging in west African countries, and results in substantial yield loss 1. TALes from Xoo are injected by a type III secretion system into plant cells and recognize effector-binding elements (EBEs) in cognate SWEET host gene promoters, which results in induction of SWEET genes and production of sugars that enable disease susceptibility in rice 2,3. An array of central repeats, which are 34-35 amino acids long, are present in each TALe and interact with EBEs via two repeat variable di-residues (RVDs) at the 12th and 13th position of each repeat 4,5. Aberrant repeats, longer than 35 amino acids, are hypothesized to allow looping out of the repeat to accommodate alternate sequence binding for a particular TALe 6. Bacterial blight depends on TALe-mediated induction of at least one member of a family of sugar-transporter genes. Although rice has more than 20 SWEET genes, only those of clade III are reported to be induced by Xoo 7-10. Although all five of the known clade III SWEET genes in rice can function as susceptibility genes for bacterial blight, only three are known to be targeted in nature 10. More specifically, SWEET11 expression is induced by strains encoding the TALe PthXo1, SWEET13 by PthXo2 and SWEET14 by any one of several TALes, namely AvrXa7, PthXo3, TalC and TalF (originally Tal5) 7,9-15 (Table 1). Effectors of Xoo that target clade III SWEET genes are referred to as major TALes owing to their strong virulence effect. Naturally occurring resistance has arisen as the result of nucleotide polymorphisms in EBEs of SWEET promoters. EBE alleles of SWEET11 that are not recognized by PthXo1 are collectively referred to as the recessive resistance gene xa13. Rice varieties containing xa13 are resistant to strains that solely depend on PthXo1 for SWEET induction. Most indica rice varieties carry a SWEET13 allele that contains four adenines in the EBE for PthXo2, and rice lines carrying this allele are susceptible to PthXo2-dependent strains 12. A rare exception is the recessive resistance allele xa25, which contains three adenines in the EBE for SWEET13 in the indica cultivar Minghui 63, conferring resistance to strains that depend solely on PthXo2 16. A similar recessive resistance allele in japonica rice varieties is equally effective against strains relying on PthXo2 (ref. 12). Additional naturally occurring recessive EBE polymorphisms that confer resistance to strains carrying PthXo2, and the overlapping EBEs for PthXo3, TalF and AvrXa7 have subsequently been identified in the promoters of SWEET13 and SWEET14, respectively, in germplasm collections 17,18. Rice susceptibility genes are good targets for genome editing for disease resistance. TALe-mediated susceptibility is particularly modifiable. For instance, transcription-activator-like effector nuclease (TALEN)-directed mutations in SWEET14 created lines resistant to strains carrying PthXo3/Avr...

Global socioeconomic developments create strong incentives for farmers to shift from transplanted to direct-seeded rice (DSR) as a means of intensification and economization(1). Rice production must increase to ensure food security(2) and the bulk of this increase will have to be achieved through intensification of cultivation, because expansion of cultivated areas is reaching sustainable limits(3). Anaerobic germination tolerance, which enables uniform germination and seedling establishment under submergence(4), is a key trait for the development of tropical DSR varieties(5,6). Here, we identify a trehalose-6-phosphate phosphatase gene, OsTPP7, as the genetic determinant in qAG-9-2, a major quantitative trait locus (QTL) for anaerobic germination tolerance(7). OsTPP7 is involved in trehalose-6-phosphate (T6P) metabolism, central to an energy sensor that determines anabolism or catabolism depending on local sucrose availability(8,9). OsTPP7 activity may increase sink strength in proliferating heterotrophic tissues by indicating low sugar availability through increased T6P turnover, thus enhancing starch mobilization to drive growth kinetics of the germinating embryo and elongating coleoptile, which consequently enhances anaerobic germination tolerance.

Increasing crop production is essential for securing the future food supply in developing countries in Asia and Africa as economies and populations grow. However, although the Green Revolution led to increased grain production in the 1960s, no major advances have been made in increasing yield potential in rice since then. In this study, we identified a gene, SPIKELET NUMBER (SPIKE), from a tropical japonica rice landrace that enhances the grain productivity of indica cultivars through pleiotropic effects on plant architecture. Map-based cloning revealed that SPIKE was identical to NARROW LEAF1 (NAL1), which has been reported to control vein pattern in leaf. Phenotypic analyses of a near-isogenic line of a popular indica cultivar, IR64, and overexpressor lines revealed increases in spikelet number, leaf size, root system, and the number of vascular bundles, indicating the enhancement of source size and translocation capacity as well as sink size. The near-isogenic line achieved 13-36% yield increase without any negative effect on grain appearance. Expression analysis revealed that the gene was expressed in all cell types: panicles, leaves, roots, and culms supporting the pleiotropic effects on plant architecture. Furthermore, SPIKE increased grain yield by 18% in the recently released indica cultivar IRRI146, and increased spikelet number in the genetic background of other popular indica cultivars. The use of SPIKE in rice breeding could contribute to food security in indica-growing regions such as South and Southeast Asia.qTSN4 | gene validation | pleiotropy | marker-assisted breeding

The homeodomain leucine zipper (HD-Zip) genes encode transcription factors that have diverse functions in plant development and have often been implicated in stress adaptation. The HD-Zip genes are the most abundant group of homeobox (HB) genes in plants and do not occur in other eukaryotes. This paper describes the complete annotation of the HD-Zip families I, II and III from rice and compares these gene families with Arabidopsis in a phylogeny reconstruction. Orthologous pairs of rice and Arabidopsis HD-Zip genes were predicted based on neighbour joining and maximum parsimony (MP) trees with support of conserved intron-exon organization. Additionally, a number of HD-Zip genes appeared to be unique to rice. Searching of EST and cDNA databases and expression analysis using RT-PCR showed that 30 out of 31 predicted rice HD-Zip genes are expressed. Most HD-Zip genes were broadly expressed in mature plants and seedlings, but others showed more organ specific patterns. Like in Arabidopsis and other dicots, a subset of the rice HD-Zip I and II genes was found to be regulated by drought stress. We identified both drought-induced and drought-repressed HD-Zip genes and demonstrate that these genes are differentially regulated in drought-sensitive versus drought-tolerant rice cultivars. The drought-repressed HD-Zip family I gene, Oshox4, was selected for promoter-GUS analysis, showing that drought-responsiveness of Oshox4 is controlled by the promoter and that Oshox4 expression is predominantly vascular-specific. Loss-of-function analysis of Oshox4 revealed no specific phenotype, but overexpression analysis suggested a role for Oshox4 in elongation and maturation processes.

More than two billion people are micronutrient deficient. Polished grains of popular rice varieties have concentration of approximately 2 μg g−1 iron (Fe) and 16 μg g−1 zinc (Zn). The HarvestPlus breeding programs for biofortified rice target 13 μg g−1 Fe and 28 μg g−1 Zn to reach approximately 30% of the estimated average requirement (EAR). Reports on engineering Fe content in rice have shown an increase up to 18 μg g−1 in glasshouse settings; in contrast, under field conditions, 4 μg g−1 was the highest reported concentration. Here, we report on selected transgenic events, field evaluated in two countries, showing 15 μg g−1 Fe and 45.7 μg g−1 Zn in polished grain. Rigorous selection was applied to 1,689 IR64 transgenic events for insert cleanliness and, trait and agronomic performances. Event NASFer-274 containing rice nicotianamine synthase (OsNAS2) and soybean ferritin (SferH-1) genes showed a single locus insertion without a yield penalty or altered grain quality. Endosperm Fe and Zn enrichment was visualized by X-ray fluorescence imaging. The Caco-2 cell assay indicated that Fe is bioavailable. No harmful heavy metals were detected in the grain. The trait remained stable in different genotype backgrounds.

Zinc (Zn) is one of the most essential micronutrients required for the growth and development of human beings. More than one billion people, particularly children and pregnant women suffer from Zn deficiency related health problems in Asia. Rice is the major staple food for Asians, but the presently grown popular high yielding rice varieties are poor supplier of Zn in their polished form. Breeding rice varieties with high grain Zn has been suggested to be a sustainable, targeted, food-based and cost effective approach in alleviating Zn deficiency. The physiological, genetic and molecular mechanisms of Zn homeostasis have been well studied, but these mechanisms need to be characterized from a biofortification perspective and should be well integrated with the breeding processes. There is a significant variation for grain Zn in rice germplasm and efforts are being directed at exploiting this variation through breeding to develop high Zn rice varieties. Several QTLs and gene specific markers have been identified for grain Zn and there is a great potential to use them in Marker-Assisted Breeding. A thorough characterization of genotype and environmental interactions is essential to identify key environmental factors influencing grain Zn. Agronomic biofortification has shown inconsistent results, but a combination of genetic and agronomic biofortification strategies may be more effective. Significant progress has been made in developing high Zn rice lines for release in target countries. A holistic breeding approach involving high Zn trait development, high Zn product development, product testing and release, including bioefficacy and bioavailability studies is essential for successful Zn biofortification.Electronic supplementary materialThe online version of this article (doi:10.1186/s12284-016-0122-5) contains supplementary material, which is available to authorized users.

SummaryRice tungro disease (RTD) is a serious constraint in rice production across tropical Asia. RTD is caused by the interaction between Rice tungro spherical virus (RTSV) and Rice tungro bacilliform virus. RTSV resistance found in traditional cultivars has contributed to a reduction in the incidence of RTD in the field. Natural RTSV resistance is a recessive trait controlled by the translation initiation factor 4 gamma gene (eIF4G). The Y1059V1060V1061 residues of eIF4G are known to be associated with the reactions to RTSV. To develop new sources of resistance to RTD, mutations in eIF4G were generated using the CRISPR/Cas9 system in the RTSV‐susceptible variety IR64, widely grown across tropical Asia. The mutation rates ranged from 36.0% to 86.6%, depending on the target site, and the mutations were successfully transmitted to the next generations. Among various mutated eIF4G alleles examined, only those resulting in in‐frame mutations in SVLFPNLAGKS residues (mainly NL), adjacent to the YVV residues, conferred resistance. Furthermore, our data suggest that eIF4G is essential for normal development, as alleles resulting in truncated eIF4G could not be maintained in homozygous state. The final products with RTSV resistance and enhanced yield under glasshouse conditions were found to no longer contain the Cas9 sequence. Hence, the RTSV‐resistant plants with the novel eIF4G alleles represent a valuable material to develop more diverse RTSV‐resistant varieties.