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Background: Deep rooting is an important factor affecting rice drought resistance. However, few genes have been identified to control this trait in rice. Previously, we identified several candidate genes by QTL mapping of the ratio of deep rooting and gene expression analysis in rice., Results: In the present work, we cloned one of these candidate genes, OsSAUR11, which encodes a small auxin-up RNA (SAUR) protein. Overexpression of OsSAUR11 significantly enhanced the ratio of deep rooting of transgenic rice, but knockout of this gene did not significantly affect deep rooting. The expression of OsSAUR11 in rice root was induced by auxin and drought, and OsSAUR11-GFP was localized both in the plasma membrane and cell nucleus. Through an electrophoretic mobility shift assay and gene expression analysis in transgenic rice, we found that the transcription factor OsbZIP62 can bind to the promoter of OsSAUR11 and promote its expression. A luciferase complementary test showed that OsSAUR11 interacts with the protein phosphatase OsPP36. Additionally, expression of several auxin synthesis and transport genes (e.g., OsYUC5 and OsPIN2) were down-regulated in OsSAUR11-overexpressing rice plants., Conclusions: This study revealed a novel gene OsSAUR11 positively regulates deep rooting in rice, which provides an empirical basis for future improvement of rice root architecture and drought resistance., (© 2023. The Author(s).)

Cold stress at the reproductive stage severely affects the production and geographic distribution of rice. The Geng/japonica subpopulation gradually developed stronger cold adaptation than the Xian/indica subpopulation during the long-term domestication of cultivated rice. However, the evolutionary path and natural alleles underlying the cold adaptability of intra-Geng subspecies remain largely unknown. Here, we identified MITOGEN-ACTIVATED PROTEIN KINASE 3 (OsMAPK3) and LATE EMBRYOGENESIS ABUNDANT PROTEIN 9 (OsLEA9) as two important regulators for the cold adaptation of Geng subspecies from a combination of transcriptome analysis and genome-wide association study. Transgenic validation showed that OsMAPK3 and OsLEA9 confer cold tolerance at the reproductive stage. Selection and evolution analysis suggested that the Geng version of OsMAPK3 (OsMAPK3 Geng ) directly evolved from Chinese Oryza rufipogon III and was largely retained in high-latitude and high-altitude regions with low temperatures during domestication. Later, the functional nucleotide polymorphism (FNP-776) in the Kunmingxiaobaigu and Lijiangxiaoheigu version of the OsLEA9 (OsLEA9 KL ) promoter originated from novel variation of intra-Geng was selected and predominantly retained in temperate Geng to improve the adaptation of Geng together with OsMAPK3 Geng to colder climatic conditions in high-latitude areas. Breeding potential analysis suggested that pyramiding of OsMAPK3 Geng and OsLEA9 KL enhanced the cold tolerance of Geng and promotes the expansion of cultivated rice to colder regions. This study not only highlights the evolutionary path taken by the cold-adaptive differentiation of intra-Geng, but also provides new genetic resources for rice molecular breeding in low-temperature areas., (© 2022 Society for Experimental Biology and John Wiley & Sons Ltd.)

Trichomes function in plant defenses against biotic and abiotic stresses; examination of glabrous lines, which lack trichomes, has revealed key aspects of trichome development and function. Tests of allelism in 51 glabrous rice (Oryza sativa) accessions collected worldwide identified OsSPL10 and OsWOX3B as regulators of trichome development in rice. Here, we report that OsSPL10 acts as a transcriptional regulator controlling trichome development. Haplotype and transient expression analyses revealed that variation in the approximately 700-bp OsSPL10 promoter region is the primary cause of the glabrous phenotype in the indica cultivar WD-17993. Disruption of OsSPL10 by genome editing decreased leaf trichome density and length in the NIL-HL6 background. Plants with genotype OsSPL10 WD-17993 /HL6 generated by crossing WD-17993 with NIL-HL6 also had fewer trichomes in the glumes. HAIRY LEAF6 (HL6) encodes another transcription factor that regulates trichome initiation and elongation, and OsSPL10 directly binds to the HL6 promoter to regulate its expression. Moreover, the transcript levels of auxin-related genes, such as OsYUCCA5 and OsPIN-FORMED1b, were altered in OsSPL10 overexpression and RNAi transgenic lines. Feeding tests using locusts (Locusta migratoria) demonstrated that non-glandular trichomes affect feeding by this herbivore. Our findings provide a molecular framework for trichome development and an ecological perspective on trichome functions., (© 2021 Institute of Botany, Chinese Academy of Sciences.)

The improvement of cold adaptation has contributed to the increased growing area of rice. Standing variation and de novo mutation are distinct natural sources of beneficial alleles in plant adaptation. However, the genetic mechanisms and evolutionary patterns underlying these sources in a single population during crop domestication remain elusive. Here we cloned the CTB2 gene, encoding a UDP-glucose sterol glucosyltransferase, for cold tolerance in rice at the booting stage. A single standing variation (I408V) in the conserved UDPGT domain of CTB2 originated from Chinese Oryza rufipogon and contributed to the cold adaptation of Oryza sativa ssp. japonica. CTB2 is located in a 56.8 kb region, including the previously reported gene CTB4a in which de novo mutation arose c. 3200 yr BP in Yunnan province, China, conferring cold tolerance. Standing variation of CTB2 and de novo mutation of CTB4a underwent stepwise selection to facilitate cold adaptation to expand rice cultivation from high-altitude to high-latitude regions. These results provide an example of stepwise selection on two kinds of variation and describe a new molecular mechanism of cold adaptation in japonica rice., (© 2021 The Authors. New Phytologist © 2021 New Phytologist Foundation.)

Genetic studies on cold tolerance at the reproductive stage in rice could lead to significant reductions in yield losses. However, knowledge about the genetic basis and adaptive differentiation, as well as the evolution and utilization of the underlying natural alleles, remains limited. Here, 580 rice accessions in two association panels were used to perform genome-wide association study, and 156 loci associated with cold tolerance at the reproductive stage were identified. Os01g0923600 and Os01g0923800 were identified as promising candidate genes in qCTB1t, a major associated locus. Through population genetic analyses, 22 and 29 divergent regions controlling cold adaptive differentiation inter-subspecies (Xian/Indica and Geng/Japonica) and intra-Geng, respectively, were identified. Joint analyses of four cloned cold-tolerance genes showed that they had different origins and utilizations under various climatic conditions. bZIP73 and OsAPX1 differentiating inter-subspecies evolved directly from wild rice, whereas the novel mutations CTB4a and Ctb1 arose in Geng during adaptation to colder climates. The cold-tolerant Geng accessions have undergone stronger selection under colder climate conditions than other accessions during the domestication and breeding processes. Additive effects of dominant allelic variants of four identified genes have been important in adaptation to cold in modern rice varieties. Therefore, this study provides valuable information for further gene discovery and pyramiding breeding to improve cold tolerance at the reproductive stage in rice., (© 2020 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Drought resistance is required in rice breeding to address the challenge of frequent droughts. However, the evolutionary mechanism of rice drought resistance is not fully understood. We investigated the genetic differentiation between upland and lowland rice domesticated in agro-ecosystems with contrasting water-soil conditions using genome-wide SNPs. We estimated morphological differences among upland and lowland rice in drought resistance and productivity through common garden experiments. Upland rice had better drought resistance but poorer productivity. The negative correlations between traits of drought resistance and productivity are attributed to the underlying genetic trade-offs through tight linkages (e.g., DCA1 and OsCesA7) or pleiotropic effects (e.g., LAX1). The genetic trade-offs are common and greatly shape the evolution of drought resistance in upland rice. In genomic regions associated with both productivity and drought resistance, signs of balancing selection were detected in upland rice, while signs of directional selection were detected in lowland rice, potentially contributing to their adaptive differentiation. Signs of balancing selection in upland rice resulted from bi-directional selection during its domestication in drought-prone upland agro-ecosystems. Using genome-wide association analysis, we identified several valuable quantitative trait loci associated with drought resistance, for which highly differentiated genes should be considered candidates. Bi-directional selection breaking tight linkages by accumulating recombination events would be applicable in breeding water-saving and drought-resistance rice., (Copyright © 2019 The Author. Published by Elsevier Inc. All rights reserved.)

Anthocyanins are naturally active substances. In this study, anthocyanins from black rice were obtained by membrane filtration and column chromatography separation. Five anthocyanin monomers in black rice extract were identified by HPLC-MS/MS, and the major anthocyanin monomer (cyanidin-3-glucoside, C3G) was purified by preparative HPLC (Pre-HPLC). The proliferative effects of the anthocyanins on Bifidobacteria and Lactobacillus were investigated by determining the media pH, bacterial populations and metabolic products. After anaerobic incubation at 37 °C for 48 h, not only the pH of the media containing C3G was lower than that of the extract of black rice anthocyanin (BRAE), but the numbers of both Bifidobacteria and Lactobacillus were also significantly increased. Furthermore, hydroxyphenylpropionic, hydroxyphenylacetic, and hydroxybenzoic acids and other metabolites were detected by GC-MS in vitro. Our results revealed that the anthocyanins and anthocyanin monomers from black rice had prebiotic activity and they were metabolized into several small molecules by Bifidobacteria and Lactobacillus.

Water-saving and drought-resistantce rice (WDR) breeding practices have greatly increased grain yield and drought resistance. To study the genetic basis of adaptation to drought, transcriptome sequences from the WDR maintainer line HuHan2B and the recurrent parent HanFengB were analyzed for alternative splicing (AS) complexity. Intron retention, the dominant AS type, accounted for 42% of the observed AS events. Differential expression analysis revealed transcripts were preferentially expressed in different varieties and conditions. Based on gene ontology predictions, the biological functions of drought-induced transcripts were significantly enriched in genes involved in transcription regulation, chloroplast components and response to abiotic stimulus in HuHan2B, whereas developmental processes for reproduction were primarily enriched in HanFengB. The regulatory network of transcription factors was driven by cohorts of transcript splicing targets, resulting in more diversified regulatory relationships due to AS complexity than in our previous findings. Moreover, several genes were validated to accumulate novel splicing transcripts in a drought-induced manner. Together, these results suggest that HuHan2B and HanFengB share similar AS features but that a subset of genes with increased levels of AS involved in transcription regulatory networks may contribute an additional level of control for genetic improvement of drought resistance in rice maintainer HuHan2B through breeding.

Plant hormones coordinate a plant's responses to environmental stimuli and the endogenous developmental programs for cell division and elongation. Brassinosteroids are among the most important of these hormones in plant development. Recently, the ubiquitin-26S-proteasome system was identified to play a key role in hormone biology. In this study, we analyzed the function of a rice (Oryza sativa) gene, DSG1, which encodes a U-box E3 ubiquitin ligase. In the dsg1 mutant (an allelic mutant of tud1), the lengths of the roots, internodes, panicles, and seeds were shorter than that in the wild-type, which was due to defects in cell division and elongation. In addition, the leaves of the dsg1 mutant were wider and curled. The DSG1 protein is nuclear- and cytoplasm-localized and does not show tissue specificity in terms of its expression, which occurs in roots, culms, leaves, sheaths, and spikelets. The dsg1 mutant is less sensitive to brassinosteroid treatment than the wild-type, and DSG1 expression is negatively regulated by brassinosteroids, ethylene, auxin, and salicylic acid. These results demonstrate that DSG1 positively regulates cell division and elongation and may be involved in multiple hormone pathways., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

Epigenetic mechanisms are crucial mediators of appropriate plant reactions to adverse environments, but their involvement in long-term adaptation is less clear. Here, we established two rice epimutation accumulation lines by applying drought conditions to 11 successive generations of two rice varieties. We took advantage of recent technical advances to examine the role of DNA methylation variations on rice adaptation to drought stress. We found that multi-generational drought improved the drought adaptability of offspring in upland fields. At single-base resolution, we discovered non-random appearance of drought-induced epimutations. Moreover, we found that a high proportion of drought-induced epimutations maintained their altered DNA methylation status in advanced generations. In addition, genes related to transgenerational epimutations directly participated in stress-responsive pathways. Analysis based on a cluster of drought-responsive genes revealed that their DNA methylation patterns were affected by multi-generational drought. These results suggested that epigenetic mechanisms play important roles in rice adaptations to upland growth conditions. Epigenetic variations have morphological, physiological and ecological consequences and are heritable across generations, suggesting that epigenetics can be considered an important regulatory mechanism in plant long-term adaptation and evolution under adverse environments.

The ongoing deficit of fresh water resource in rice growing regions has made the selection of water-saving and drought-resistance rice (WDR) a crucial factor in developing sustainable cultivation. HuHan2B, a new japonica maintainer for WDR breeding, had the same yield potential as recurrent parent HanFengB but showed improved drought resistance in fields. We investigated the genomic content accumulation and candidate genes passed from parent to offspring using the genomic and transcriptomic approaches. The genomic constitution indicated that the genetic similarity was 84% between HuHan2B and HanFengB; additionally, 7,256 genes with specific alleles were inherited by HuHan2B from parents other than HanFengB. The differentially expressed genes (DEGs) under drought stress showed that biological function was significantly enriched for transcript regulation in HuHan2B, while the oxidation-reduction process was primarily enriched in HanFengB. Furthermore, 36 DEGs with specific inherited alleles in HuHan2B were almost involved in the regulatory network of TFs and target genes. These findings suggested that major-effect genes were congregated and transformed into offspring in manner of interacting network by breeding. Thus, exploiting the potential biological function of allelic-influencing DEGs would be of great importance for improving selection efficiency and the overall genetic gain of multiple complex traits.

Deep rooting is a very important trait for plants' drought avoidance, and it is usually represented by the ratio of deep rooting (RDR). Three sets of rice populations were used to determine the genetic base for RDR. A linkage mapping population with 180 recombinant inbred lines and an association mapping population containing 237 rice varieties were used to identify genes linked to RDR. Six quantitative trait loci (QTLs) of RDR were identified as being located on chromosomes 1, 2, 4, 7, and 10. Using 1 019 883 single-nucleotide polymorphisms (SNPs), a genome-wide association study of the RDR was performed. Forty-eight significant SNPs of the RDR were identified and formed a clear peak on the short arm of chromosome 1 in a Manhattan plot. Compared with the shallow-rooting group and the whole collection, the deep-rooting group had selective sweep regions on chromosomes 1 and 2, especially in the major QTL region on chromosome 2. Seven of the nine candidate SNPs identified by association mapping were verified in two RDR extreme groups. The findings from this study will be beneficial to rice drought-resistance research and breeding., (© The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

A recombinant inbred line (RIL) population from a cross between 'HH1B' and 'RSB02' (a deep-water rice variety with resistance to sheath blight) was planted in two locations for four different growing seasons. Seven traits were used to evaluate the disease severity, namely disease rating (DR), lesion length (LL), lesion height (LH), relative lesion length (RLL), relative lesion height (RLH), plant height (PH) and heading date (HD). Based on a linkage map of 163 simple sequence repeat (SSR) markers, a total of 37 QTLs were mapped on nine chromosomes. Additionally, 32 epistatic QTLs were identified, distributed on all the 12 chromosomes. The contribution of a single QTL's additive and epistatic effect was of low magnitude for most cases (from 0.39% to 24.62%). Among QTL × environment interaction test, 28 additive QTLs and six pairs of epistatic interactions were involved. Correlation analysis showed that DR had significant positive correlations with LL, RLL and RLH, but had a negative correlation with PH, two of six QTLs controlling DR were mapped in the same chromosome regions as the QTLs controlling PH. The alleles which can enhance disease resistance and increase PH are from the resistant parent 'RSB02', indicating that PH has certain effect on sheath blight resistance in the present study., (© 2014 The Authors.)

Adverse environmental conditions have large impacts on plant growth and crop production. One of the crucial mechanisms that plants use in variable and stressful natural environments is gene expression modulation through epigenetic modification. In this study, two rice varieties with different drought resistance levels were cultivated under drought stress from tilling stage to seed filling stage for six successive generations. The variations in DNA methylation of the original generation (G0) and the sixth generation (G6) of these two varieties in normal condition (CK) and under drought stress (DT) at seedling stage were assessed by using Methylation Sensitive Amplification Polymorphism (MSAP) method. The results revealed that drought stress had a cumulative effect on the DNA methylation pattern of both varieties, but these two varieties had different responses to drought stress in DNA methylation. The DNA methylation levels of II-32B (sensitive) and Huhan-3 (resistant) were around 39% and 32%, respectively. Genome-wide DNA methylation variations among generations or treatments accounted for around 13.1% of total MSAP loci in II-32B, but was only approximately 1.3% in Huhan-3. In II-32B, 27.6% of total differentially methylated loci (DML) were directly induced by drought stress and 3.2% of total DML stably transmitted their changed DNA methylation status to the next generation. In Huhan-3, the numbers were 48.8% and 29.8%, respectively. Therefore, entrainment had greater effect on Huhan-3 than on II-32B. Sequence analysis revealed that the DML were widely distributed on all 12 rice chromosomes and that it mainly occurred on the gene's promoter and exon region. Some genes with DML respond to environmental stresses. The inheritance of epigenetic variations induced by drought stress may provide a new way to develop drought resistant rice varieties.

Water status is the main factor affecting rice production. In order to understand rice strategies in response to drought condition in the field, the drought-responsive mechanisms at the physiological and molecular levels were studied in two rice genotypes with contrasting susceptibility to drought stress at reproductive stage. After 20 d of drought treatment, the osmotic potential of leaves reduced 78% and 8% in drought susceptible rice cultivar Zhenshan97B and tolerant rice cultivar IRAT109, respectively. The panicle lengths had no obvious changes in drought stressed Zhenshan97B and IRAT109, suggesting that drought stress impose less effect on assimilate translocation from leaf to vegetative growth of panicles. IRAT109 showed more extensive deeper root growth that could be considered a second line of defense against drought stress. The C(i)/C(a) ratio exhibited enhancement over reduction of g(s) in both cultivars, reflecting the non-stomatal limitation to photosynthesis occurred during drought stress. Orthophosphate dikinase, glycine dehydrogenase, ribulose bisphosphate carboxylase (Rubisco), glycine hydroxymethyltransferase and ATP synthase were down-regulated for Zhenshan97B in response to drought stress, suggesting the reduction of capacity of carbon assimilation in this rice cultivar. In drought-stressed IRAT109, transketolase, Rubisco were down-regulated, however, Rubisco activase and peptidyl-prolyl cis-trans isomerase, which might alleviate the damage on Rubisco by drought stress, were up-regulated. The increased abundances of chloroplastic superoxide dismutase [Cu-Zn] and dehydroascorbate reductase might provide antioxidant protection for IRAT109 against damage by dehydration., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

We used proteomic analysis to determine the response of rice plant seedlings to drought-induced stress. The expression of 71 protein spots was significantly altered, and 60 spots were successfully identified. The greatest down-regulated protein functional category was translation. Up-regulated proteins were mainly related to protein folding and assembly. Additionally, many proteins involved in metabolism (e.g. carbohydrate metabolism) also showed differences in expression. cDNA microarray and GC-MS analysis showed 4756 differentially expressed mRNAs and 37 differentially expressed metabolites. Once these data were integrated with the proteomic analysis, we were able to elucidate the metabolic pathways affected by drought-induced stress. These results suggest that increased energy consumption from storage substances occurred during drought. In addition, increased expression of the enzymes involved in anabolic pathways corresponded with an increase in the content of six amino acids. We speculated that energy conversion from carbohydrates and/or fatty acids to amino acids was increased. Analysis of basic metabolism networks allowed us to understand how rice plants adjust to drought conditions., (Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

In this study, metabolic profiles of a set of 48 rice germplasms from the Chinese core collection were obtained by gas chromatography and time-of-flight mass spectrometry (GC-TOF-MS). Forty-one metabolites were identified and relatively quantified according to the internal standard (IS). Wide ranges of variations for all metabolites were observed among rice accessions. The maximum/minimum ratios varied from 4.73 to 211.36. The metabolites were categorized into seven groups based on their chemical characteristics. Clustering analysis and a correlation network showed that most of the metabolites had variations among rice accessions in the same direction. Using 218 molecular markers, association mapping was conducted to identify the chromosomal loci influencing the concentrations of identified metabolites. Twenty markers were identified associating with the concentrations of 29 metabolites [-lg(P) > 3]. Allelic effects were investigated in detail in two markers (RM315 and RM541) as examples.