Wheat plant height locus RHT25 encodes a PLATZ transcription factor that interacts with DELLA (RHT1).

Plant height is an important agronomic trait with a significant impact on grain yield, as demonstrated by the positive effect of the REDUCED HEIGHT (RHT) dwarfing alleles (Rht1b) on lodging and harvest index in the "Green Revolution" wheat varieties. However, these gibberellic acid (GA)-insensitive alleles also reduce coleoptile length, biomass production, and yield potential in some environments, triggering the search for alternative GA-sensitive dwarfing genes. Here we report the identification, validation, and characterization of the gene underlying the GA-sensitive dwarfing locus RHT25 in wheat. This gene, designated as PLATZ-A1 (TraesCS6A02G156600), is expressed mainly in the elongating stem and developing spike and encodes a plant-specific AT-rich sequence- and zinc-binding protein (PLATZ). Natural and induced loss-of-function mutations in PLATZ-A1 reduce plant height and its overexpression increases plant height, demonstrating that PLATZ-A1 is the causative gene of RHT25. PLATZ-A1 and RHT1 show a significant genetic interaction on plant height, and their encoded proteins interact with each other in yeast and wheat protoplasts. These results suggest that PLATZ1 can modulate the effect of DELLA on wheat plant height. We identified four natural truncation mutations and one promoter insertion in PLATZ-A1 that are more frequent in modern varieties than in landraces, suggesting positive selection during wheat breeding. These mutations can be used to fine-tune wheat plant height and, in combination with other GA-sensitive dwarfing genes, to replace the GA-insensitive Rht1b alleles and search for grain yield improvements beyond those of the Green Revolution varieties.

Phenotypic and transcriptomic characterization of a wheat tall mutant carrying an induced mutation in the C-terminal PFYRE motif of RHT-B1b.

BACKGROUND: As central regulators of the gibberellic acid (GA) signaling pathway in plants, DELLA proteins function as growth repressors and affect diverse biological processes. The wheat RHT-B1b and RHT-D1b semi-dwarfing alleles, which encode GA-insensitive DELLA proteins, have been widely adopted in modern wheat varieties to improve lodging tolerance and harvest index. However, the molecular mechanisms by which DELLA modulates these responses in wheat remain largely unknown. RESULTS: We identified a tall tetraploid wheat mutant line carrying an induced missense mutation (E529K) in the PFYRE motif of RHT-B1b that partially suppressed the semi-dwarf phenotype. The height-increasing effect of RHT-B1bE529K relative to RHT-B1b (19 cm or 21% increase) was significantly smaller than the effect of RHT-B1a (33 cm or 34% increase) relative to RHT-B1b in the same field experiment. The RHT-B1bE529K mutation was also associated with length increases in coleoptiles, seedling shoots, and stem internodes relative to the RHT-B1b allele. We detected no significant differences in germination rate, seedling root length, tiller number, flag leaf size, spike length, or yield components. Using RNA-seq, we compared gene expression profiles of plants encoding RHT-B1b and RHT-B1bE529K in coleoptile, first leaf, and elongating peduncles. We detected limited overlap among tissues of the genes differentially regulated by the two genotypes, and more genes upregulated (77%) than downregulated (23%) in RHT-B1bE529K relative to RHT-B1b. These results suggest that the wheat DELLA protein affects the transcriptome in a tissue-specific manner and that the mutation mainly eliminates or reduces repression functions of the RHT-B1 protein. Our study identified distinct sets of potential DELLA direct or indirect target genes involved in cell wall and carbohydrate metabolisms, cell cycle/division, and hormone pathways. CONCLUSIONS: We identified the hypomorphic RHT-B1bE529K allele that confers an intermediate plant height and coleoptile elongation. This allele can be useful in rain-fed wheat breeding programs where the strong reduction in height and biomass associated with RHT-B1b has detrimental effects. Transcriptomic characterization of different tissues from the plants encoding RHT-B1bE529K and RHT-B1b provided valuable information for identifying DELLA downstream GA response genes in wheat.

Mapping causal mutations by exome sequencing in a wheat TILLING population: a tall mutant case study.

Forward genetic screens of induced mutant plant populations are powerful tools to identify genes underlying phenotypes of interest. Using traditional techniques, mapping causative mutations from forward screens is a lengthy, multi-step process, requiring the identification of a broad genetic region followed by candidate gene sequencing to characterize the causal variant. Mapping by whole genome sequencing accelerates the identification of causal mutations by simultaneously defining a mapping region and providing information on the induced genetic variants. In wheat, although the availability of a high-quality draft genome assembly facilitates mapping and mutation calling, whole genome resequencing remains prohibitively expensive due to its large genome. In the current study, we used exome sequencing as a complexity reduction strategy to detect mutations associated with a target phenotype. In a segregating wheat EMS population, we identified a clear peak region on chromosome arm 4BS associated with increased plant height. Although none of the significant SNPs seemed causative for the mutant phenotype, they were sufficient to identify a linked ~ 1.9 Mb deletion encompassing nine genes. These genes included Rht-B1, which is known to have a strong effect on plant height and is a strong candidate for the observed phenotype. We performed simulation experiments to determine the impacts of sequencing depth and bulk size and discuss the importance of considering each factor when designing mapping-by-sequencing experiments in wheat. This approach can accelerate the identification of candidate causal point mutations or linked deletions underlying important phenotypes.

A new SNP in cyOsPPDK gene is associated with floury endosperm in Suweon 542.

Pyruvate orthophosphate dikinase (PPDK) is a component of glycolysis to mediate endosperm energy charge by adjusting the ratio of ATP to ADP and AMP that proposed to balance the flow of carbon into starch, protein, fatty acid and amino acid biosynthesis. However, these were inconsistent with the first report of a T-DNA insertional knockout mutant of the rice PPDK gene (flo4) showed that rice with inactivated PPDK gene failed to produce a opaque seeds. Therefore, the PPDK might have multifaceted functions in grain filling stage, which in some ways might depend on the direction of the reversible catalysis. Suweon 542 is a rice (Oryza sativa L.) mutant developed from Oryza sativa ssp. japonica cv. Namil. Suweon 542 has a milky-white floury endosperm suitable for dry filling, with low starch damage, low grain hardness, and fine flour particle size. The mutant locus on chromosome 5 controls the floury endosperm phenotype of Suweon 542. Fine mapping of this locus is required for efficient breeding of rice germplasm suitable for dry milling. In this study, whole genome of Suweon 542 and Milyang 23 were re-sequenced using Illumina HiSeq 2500. Co-segregation analysis of F3:4 family populations derived from Suweon 542/Milyang 23 was performed using eight CAPS markers and phenotypic evaluation of the endosperm. The target region was mapped to a 33 kb region and identified to encode cytosolic pyruvate orthophosphate dikinase protein (cyOsPPDK). A G→A SNP in exon 8 of cyOsPPDK resulting in a missense mutation from Gly to Asp at amino acid position 404 was responsible for the floury endosperm of Suweon 542. qRT-PCR experiments revealed that FLO4-4 was expressed to a considerably higher level in Suweon 542 than in Namil during the grain filling stage. Overall, fine mapping of FLO4-4 and candidate gene analysis provided further insight into the floury endosperm of rice, and reveal a novel SNP in cyOsPPDK gene can affect the floury endosperm phenotype through active PPDK gene during grain filling stage.

Identification and characterization of Rht25, a locus on chromosome arm 6AS affecting wheat plant height, heading time, and spike development.

KEY MESSAGE: This study identified Rht25, a new plant height locus on wheat chromosome arm 6AS, and characterized its pleiotropic effects on important agronomic traits. Understanding genes regulating wheat plant height is important to optimize harvest index and maximize grain yield. In modern wheat varieties grown under high-input conditions, the gibberellin-insensitive semi-dwarfing alleles Rht-B1b and Rht-D1b have been used extensively to confer lodging tolerance and improve harvest index. However, negative pleiotropic effects of these alleles (e.g., poor seedling emergence and reduced biomass) can cause yield losses in hot and dry environments. As part of current efforts to diversify the dwarfing alleles used in wheat breeding, we identified a quantitative trait locus (QHt.ucw-6AS) affecting plant height in the proximal region of chromosome arm 6AS (< 0.4 cM from the centromere). Using a large segregating population (~ 2800 gametes) and extensive progeny tests (70-93 plants per recombinant family), we mapped QHt.ucw-6AS as a Mendelian locus to a 0.2 cM interval (144.0-148.3 Mb, IWGSC Ref Seq v1.0) and show that it is different from Rht18. QHt.ucw-6AS is officially designated as Rht25, with Rht25a representing the height-increasing allele and Rht25b the dwarfing allele. The average dwarfing effect of Rht25b was found to be approximately half of the effect observed for Rht-B1b and Rht-D1b, and the effect is greater in the presence of the height-increasing Rht-B1a and Rht-D1a alleles than in the presence of the dwarfing alleles. Rht25b is gibberellin-sensitive and shows significant pleiotropic effects on coleoptile length, heading date, spike length, spikelet number, spikelet density, and grain weight. Rht25 represents a new alternative dwarfing locus that should be evaluated for its potential to improve wheat yield in different environments.

Effects of allelic variations in starch synthesis-related genes on grain quality traits of Korean nonglutinous rice varieties under different temperature conditions.

Influences of allelic variations in starch synthesis-related genes (SSRGs) on rice grain quality were examined. A total of 187 nonglutinous Korean rice varieties, consisting of 170 Japonica and 17 Tongil-type varieties, were grown in the field and in two greenhouse conditions. The percentages of head rice and chalky grains, amylose content, alkali digestion value, and rapid visco-analysis characteristics were evaluated in the three different environments. Among the 10 previously reported SSRG markers used in this study, seven were polymorphic, and four of those showed subspecies-specific allele distributions. Six out of the seven polymorphic SSRG markers were significantly associated with at least one grain quality trait (R (2) > 0.1) across the three different environments. However, the association level and significance were markedly lower when the analysis was repeated using only the 170 Japonica varieties. Similarly, the significant associations between SSRG allelic variations and changes in grain quality traits under increased temperature were largely attributable to the biased allele frequency between the two subpopulations. Our results suggest that within Korean Japonica varieties, these 10 major SSRG loci have been highly fixed during breeding history and variations in grain quality traits might be influenced by other genetic factors.

Allelic combinations of Hd1, Hd16, and Ghd7 exhibit pleiotropic effects on agronomic traits in rice.

Heading date is a critical agronomic trait that significantly affects grain yield and quality in rice. As early heading is typically associated with reduced yield due to shorter growth duration, it is essential to harness optimum heading date genes and their allelic combinations to promote heading while minimizing yield penalties. In this study, we identified quantitative trait loci (QTLs) for heading date and other major agronomic traits in a recombinant inbred line (RIL) population derived from a cross between Koshihikari and Baegilmi. Analyses on 3 major QTLs for heading date and their underlying genes (Hd1, Hd16, and Ghd7) revealed their pleiotropic effects on culm length, panicle length, and head rice percentage. Additionally, Ghd7 exhibited pleiotropic effects on panicle number and grain size. Among 8 different types of allelic combinations of the 3 heading date genes, RILs carrying a single nonfunctional hd16 or ghd7 under the functional background of the other 2 genes (Hd1hd16Ghd7 and Hd1Hd16ghd7) showed potential for maintaining yield and quality-related traits while accelerating heading. These results provide valuable insights for fine-tuning heading dates in rice breeding programs.

GWAS analysis reveals the genetic basis of blast resistance associated with heading date in rice.

Rice blast is a destructive fungal disease affecting rice plants at various growth stages, significantly threatening global yield stability. Development of resistant rice cultivars stands as a practical means of disease control. Generally, association mapping with a diversity panel powerfully identifies new alleles controlling trait of interest. On the other hand, utilization of a breeding panel has its advantage that can be directly applied in a breeding program. In this study, we conducted a genome-wide association study (GWAS) for blast resistance using 296 commercial rice cultivars with low population structure but large phenotypic diversity. We attempt to answer the genetic basis behind rice blast resistance among early maturing cultivars by subdividing the population based on its Heading date 1 (Hd1) functionality. Subpopulation-specific GWAS using the mixed linear model (MLM) based on blast nursery screening conducted in three years revealed a total of 26 significant signals, including three nucleotide-binding site leucine-rich repeat (NBS-LRR) genes (Os06g0286500, Os06g0286700, and Os06g0287500) located at Piz locus on chromosome 6, and one at the Pi-ta locus (Os12g0281300) on chromosome 12. Haplotype analysis revealed blast resistance associated with Piz locus was exclusively specific to Type 14 hd1 among japonica rice. Our findings provide valuable insights for breeding blast resistant rice and highlight the applicability of our elite cultivar panel to detect superior alleles associated with important agronomic traits.

Insights into chloroplast genome structure and phylogenetic relationships within the Sesamum species complex (Pedaliaceae).

Background: In the Sesamum species complex, the lack of wild species genomic resources hinders the evolutionary comprehension of phylogenetic relationships. Results: In the present study, we generated complete chloroplast genomes of six wild relatives (Sesamum alatum, Sesamum angolense, Sesamum pedaloides, Ceratotheca sesamoides (syn. Sesamum sesamoides), Ceratotheca triloba (syn. Sesamum trilobum), and Sesamum radiatum) and a Korean cultivar, Sesamum indicum cv. Goenbaek. A typical quadripartite chloroplast structure, including two inverted repeats (IR), a large single copy (LSC), and a small single copy (SSC), was observed. A total of 114 unique genes encompassing 80 coding genes, four ribosomal RNAs, and 30 transfer RNAs were counted. The chloroplast genomes (152, 863-153, 338 bp) exhibited the IR contraction/expansion phenomenon and were quite conserved in both coding and non-coding regions. However, high values of the nucleotide diversity index were found in several genes, including ndhA, ndhE, ndhF, ycf1, and psaC-ndhD. Concordant tree topologies suggest ndhF as a useful marker for taxon discrimination. The phylogenetic inference and time divergence dating indicate that S. radiatum (2n = 64) occurred concomitantly with the sister species C. sesamoides (2n = 32) approximately 0.05 million years ago (Mya). In addition, S. alatum was clearly discriminated by forming a single clade, showing its long genetic distance and potential early speciation event in regards to the others. Conclusion: Altogether, we propose to rename C. sesamoides and C. triloba as S. sesamoides and S. trilobum, respectively, as suggested previously based on the morphological description. This study provides the first insight into the phylogenetic relationships among the cultivated and wild African native relatives. The chloroplast genome data lay a foundation for speciation genomics in the Sesamum species complex.

Characterization of Peroxidase and Laccase Gene Families and In Silico Identification of Potential Genes Involved in Upstream Steps of Lignan Formation in Sesame.

Peroxidases and laccases are oxidative enzymes involved in physiological processes in plants, covering responses to biotic and abiotic stress as well as biosynthesis of health-promoting specialized metabolites. Although they are thought to be involved in the biosynthesis of (+)-pinoresinol, a comprehensive investigation of this class of enzymes has not yet been conducted in the emerging oil crop sesame and no information is available regarding the potential (+)-pinoresinol synthase genes in this crop. In the present study, we conducted a pan-genome-wide identification of peroxidase and laccase genes coupled with transcriptome profiling of diverse sesame varieties. A total of 83 and 48 genes have been identified as coding for sesame peroxidase and laccase genes, respectively. Based on their protein domain and Arabidopsis thaliana genes used as baits, the genes were classified into nine and seven groups of peroxidase and laccase genes, respectively. The expression of the genes was evaluated using dynamic transcriptome sequencing data from six sesame varieties, including one elite cultivar, white vs black seed varieties, and high vs low oil content varieties. Two peroxidase genes (SiPOD52 and SiPOD63) and two laccase genes (SiLAC1 and SiLAC39), well conserved within the sesame pan-genome and exhibiting consistent expression patterns within sesame varieties matching the kinetic of (+)-pinoresinol accumulation in seeds, were identified as potential (+)-pinoresinol synthase genes. Cis-acting elements of the candidate genes revealed their potential involvement in development, hormonal signaling, and response to light and other abiotic triggers. Transcription factor enrichment analysis of promoter regions showed the predominance of MYB binding sequences. The findings from this study pave the way for lignans-oriented engineering of sesame with wide potential applications in food, health and medicinal domains.

Novel Sources of Pre-Harvest Sprouting Resistance for Japonica Rice Improvement.

Pre-harvest sprouting (PHS), induced by unexpected weather events, such as typhoons, at the late seed maturity stage, is becoming a serious threat to rice production, especially in the state of California, USA, Japan, and the Republic of Korea, where japonica varieties (mostly susceptible to PHS) are mainly cultivated. A projected economic loss by severe PHS in these three countries could range between 8-10 billion USD per year during the next 10 years. Here, we present promising rice germplasm with strong resistance to PHS that were selected from a diverse rice panel of accessions held in the International Rice Genebank (IRG) at the International Rice Research Institute (IRRI). To induce PHS, three panicle samples per accession were harvested at 20 and 30 days after flowering (DAF), respectively, and incubated at 100% relative humidity (RH), 30 °C in a growth chamber for 15 days. A genome-wide association (GWA) analysis using a 4.8 million single nucleotide polymorphisms (SNP) marker set was performed to identify loci and candidate genes conferring PHS resistance. Interestingly, two tropical japonica and four temperate japonica accessions showed outstanding PHS resistance as compared to tolerant indica accessions. Two major loci on chromosomes 1 and 4 were associated with PHS resistance. A priori candidate genes interactions with rice gene networks, which are based on the gene ontology (GO), co-expression, and other evidence, suggested that a key resistance mechanism is related to abscisic acid (ABA), gibberellic acid (GA), and auxin mediated signaling pathways.

Hd1 Allele Types and Their Associations with Major Agronomic Traits in Korean Rice Cultivars.

Optimizing flowering time in crop plants is critical for maximizing yield and quality under target environments. While there is a wide range of heading date variation in Korean rice cultivars, the underlying gene mechanisms are unclear. Here, we sequenced the protein coding regions of Hd1, the major rice heading date gene, from 293 Korean rice cultivars and investigated the associations between Hd1 allele types and major agronomic traits under four different environments. There were four functional Hd1 and five nonfunctional hd1 alleles distributed among the 293 Korean rice cultivars. The effects of the Hd1 allele types were highly significant for days to heading in all four environments, explaining 51.4-65.8% of the phenotypic variation. On average, cultivars carrying nonfunctional hd1 headed 13.7 days earlier than those carrying functional Hd1. While the Hd1 allele types exhibited highly significant effects on culm length and protein content under all four environments, the differences between cultivars carrying Hd1 and hd1 were minimal. The effects of the Hd1 allele types on amylose content were significant in only one of the four environments. Our results provide useful information for fine-tuning rice heading dates by utilizing different Hd1 alleles in rice breeding programs.

Characterization of QTLs and Candidate Genes for Days to Heading in Rice Recombinant Inbred Lines.

Understanding the gene mechanisms controlling days to heading (DH) is important in rice breeding for adaption in the target environment. Using a recombinant inbred line population derived from the cross between two japonica rice cultivars, Koshihikari and Baegilmi, we identified three consistent quantitative trait loci (QTLs) for DH for two years, qDH3, qDH6, and qDH7 on chromosomes 3, 6, and 7, respectively. While Baegilmi contributed the allele for early heading at qDH6 and qDH7 with the additive effect of five days each, Koshihikari contributed the allele for early heading at qDH3 with the additive effect of three days. Notably, pyramiding two or more alleles for early heading at these QTLs accelerated heading effectively. Sequencing of Hd16, Hd1, and Ghd7, the previously known heading date genes underlying qDH3, qDH6, and qDH7, respectively, revealed that Baegilmi and Koshihikari carry different alleles at the three genes. Molecular markers were developed to screen the allelic compositions of the three genes among 295 Korean commercial rice cultivars. The results showed that few cultivars carry alleles for early heading at the three genes, highlighting that DH can be further accelerated and fine-tuned in breeding programs by combining the desirable alleles of Hd16, Hd1, and Ghd7.

A New SNP in Rice Gene Encoding Pyruvate Phosphate Dikinase (PPDK) Associated with Floury Endosperm.

Rice varieties with suitable flour-making qualities are required to promote the rice processed-food industry and to boost rice consumption. A rice mutation, Namil(SA)-flo1, produces grains with floury endosperm. Overall, grains with low grain hardness, low starch damage, and fine particle size are more suitable for use in flour processing grains with waxy, dull endosperm with normal grain hardness and a high amylose content. In this study, fine mapping found a C to T single nucleotide polymorphism (SNP) in exon 2 of the gene encoding cytosolic pyruvate phosphate dikinase (cyOsPPDK). The SNP resulted in a change of serine to phenylalanine acid at amino acid position 101. The gene was named FLOURY ENDOSPERM 4-5 (FLO4-5). Co-segregation analysis with the developed cleaved amplified polymorphic sequence (CAPS) markers revealed co-segregation between the floury phenotype and the flo4-5. This CAPS marker could be applied directly for marker-assisted selection. Real-time RT-PCR experiments revealed that PPDK was expressed at considerably higher levels in the flo4-5 mutant than in the wild type during the grain filling stage. Plastid ADP-glucose pyrophosphorylase small subunit (AGPS2a and AGPS2b) and soluble starch synthase (SSIIb and SSIIc) also exhibited enhanced expression in the flo4-5 mutant.

Agronomic and genetic analysis of Suweon 542, a rice floury mutant line suitable for dry milling.

BACKGROUND: Producing rice flour of good quality by dry milling is necessary to reduce milling costs and promote the processed rice food industry. This study was conducted to evaluate the dry milling properties of Suweon 542, a floury endosperm mutant, and identify the chromosomal region responsible for the floury endosperm characteristics. RESULTS: Compared with the wild type, after dry milling process, the grain hardness of Suweon 542 was significantly lower because of its round and loosely packed starch granules. Also, the flour of Suweon 542 had significantly smaller particles and less damaged starch than Namil and other rice cultivars and its particle size distribution was similar to a commercial wheat cultivar. Considering that the yield loss of Suweon 542 due to its floury endosperm was largely compensated for by an increased number of spikelets per panicle, Suweon 542 has potential value as a raw material for rice flour production. Association analysis using 70 genome-wide SSR markers and 94 F2 plants derived from Suweon 542/Milyang 23 showed that markers on chromosome 5 explained a large portion of the variation in floury grains percentage (FGP). Further analysis with an increased number of SSR markers revealed that the floury endosperm of Suweon 542 was directed by a major recessive locus, flo7(t), located in the 19.33-19.86 Mbp region of chromosome 5, with RM18639 explaining 92.2% of FGP variation in the F2 population. CONCLUSIONS: The floury endosperm of Suweon 542 is suitable for dry milling, with a small flour particle size and low damaged starch content. Further physical mapping of flo7(t), the floury endosperm locus of Suweon 542, would facilitate efficient breeding of rice cultivars with proper dry milling adaptability that can be used in the processed rice food industry.