Background/Objectives: Having longer mesocotyls is beneficial for the deep-sowing tolerance of rice, which is important for seedling establishment. Methods: Here, we performed transcriptome analysis of the elongating mesocotyl of Zhengdao 209 in response to three different sowing depths to identify the pivotal genes regulating rice mesocotyl elongation. Results: Three groups with different mesocotyl lengths were compared using transcriptome analysis, and 60 common differentially expressed genes were detected. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses revealed that these genes are primarily involved in phenylpropanoid biosynthesis, cutin suberine and wax biosynthesis, the plant mitogen-activated protein kinase signaling pathway, diterpenoid biosynthesis, cyanoamino acid metabolism, carbon fixation in photosynthetic organisms, flavonoid biosynthesis, and glutathione metabolism. Furthermore, weighted gene co-expression network and hierarchical clustering analyses showed that most of the differentially expressed genes are implicated in phenylpropanoid biosynthesis, carbon metabolism, photosynthesis antenna proteins, and plant-pathogen interactions. Among the genes involved in phenylpropanoid biosynthesis processes, the expression levels of OsPHT3 and LOC_Os04g59260 increased, while OsCCR1, OsPGIP4, and LOC_Os01g45110 expression decreased with increasing sowing depth. Among the genes involved in the mitogen-activated protein kinase signaling pathway, the expression levels of LOC_Os07g03319 and LOC_Os07g03580 increased, while LOC_Os07g03409 decreased with increasing sowing depth. Among the genes involved in diterpenoid biosynthesis processes, the expression levels of OsCYP76M5 and OsCYP71Z2 decreased, while OsCYP71Z21 increased with increasing sowing depth. Furthermore, the expression levels of these genes were analyzed using quantitative real-time polymerase chain reaction, which confirmed the transcriptome analysis results. Conclusions: This study identified candidate genes governing rice mesocotyl length and provides novel insights into the molecular regulatory mechanisms underlying mesocotyl elongation in rice.

Understanding the regulatory mechanisms underlying the plant heat stress response is important for developing climate-resilient crops, including rice (Oryza sativa). Here, we report that OsEIL5, one member of the ETHYLENE INSENSITIVE3-LIKE family, positively regulates rice heat tolerance at the seedling and reproductive stages. OsEIL5 directly binds to the promoter of OsPP91 (encoding one type 2C protein phosphatase) and activates its expression. OsPP91 is required for rice thermotolerance, and overexpressing OsPP91 in oseil5-1 partially rescues its heat sensitivity. The F box protein OsEBF1 interacts with OsEIL5 and degrades it through the ubiquitination pathway, resulting in the reduction of OsPP91 expression and ultimately weakening rice heat tolerance. Knocking out OsEIL5 in the EBF1R13 line partially reduces its extremely high heat tolerance. Taken together, our work uncovers a mechanism that finely regulates rice thermotolerance through the OsEBF1-OsEIL5-OsPP91 module at the posttranslational and transcriptional levels.

Global warming and uneven distribution of rainfall have led to frequent drought events. Drought is a major obstacle affecting rice production; therefore, it is crucial to elucidate the drought regulatory mechanisms and breed drought-resistant and water-saving rice varieties. With the rapid development of molecular biology tools, numerous drought-related genes have been identified and cloned. At present, numerous genes related to osmotic regulation, antioxidant scavenging, phytohormones, root development, stoma and cuticle regulation have been found to improve drought resistance. This review summarizes the research progress in the cloning of drought tolerance and avoidance genes in rice, their functional characterization and validation. It reveals that current studies on drought-resistance molecular pathways and the identification of drought-resistant genes remain limited. More critically, only a few genes have been tested for drought resistance under field conditions and shown to enhance both drought resistance and yield in field environments. In the future, greater emphasis should be placed on discovering drought-resistant genes from upland rice resources. Additionally, rigorous testing and evaluation of these genes under field drought conditions are essential to assess their breeding potential, thereby advancing molecular breeding for drought resistance.

Submergence stress is a major obstacle limiting the application of direct seeding in rice cultivation. Therefore, understanding the genetic basis of submergence tolerance in rice is of great significance for identifying favorable genes and developing superior rice varieties. However, few studies have focused on submergence tolerance during seed germination; thus, the genetic basis of submergence tolerance at this stage deserves more attention. In this study, a natural population of 432 rice varieties collected from 25 provinces in China and other countries was used for the first time to evaluate submergence tolerance during seed germination. Population structure analysis revealed that this population was divided into two groups and was rich in genetic diversity. Our findings confirm that japonica rice is more tolerant to submergence than indica rice during seed germination. A genome-wide association study (GWAS) was performed using the phenotypic data of 432 rice varieties and a 3,548,101 single-nucleotide polymorphism (SNP) dataset using a mixed linear model. Thirteen QTLs (P < 0.0001) were identified for the coleoptile length (CL) under submergence (10 cm depth of water), of which four QTLs (qCL3-1, qCL4-2, qCL5-1, and qCL5-2) colocalized with those from previous studies, while nine QTLs (qCL2-1, qCL2-2, qCL2-3, qCL2-4, qCL3-2, qCL4-1, qCL8-1, qCL8-2, and qCL9) were reported for the first time. Among these QTLs, qCL9, which harbored the most significant SNP, explained most of the phenotypic variation. Using quantitative real-time polymerase chain reaction (qRT-PCR) analysis and phenotypic identification of the knockout lines, three genes (LOC_Os09g11590, LOC_Os09g11660, and LOC_Os09g11760) were identified as candidates for qCL9. There were 28, 13, and 13 non-synonymous SNPs in LOC_Os09g11590, LOC_Os09g11660, and LOC_Os09g11760, respectively, from which 5, 3, and 5 haplotypes were detected in rice varieties. This natural population, consisting of 432 rice varieties combined with high-density SNPs, provides a valuable resource for identifying rice QTLs/genes in the future. The detected QTLs/genes associated with submergence tolerance during seed germination in rice offer new insights for gene discovery and will facilitate the breeding of rice varieties with improved tolerance to submergence stress.

Heat stress significantly impacts global rice production, highlighting the critical need to understand the genetic basis of heat resistance in rice. U2AF (U2 snRNP auxiliary factor) is an essential splicing complex with critical roles in recognizing the 3'-splice site of precursor messenger RNAs (pre-mRNAs). The U2AF small subunit (U2AF35) can bind to the 3'-AG intron border and promote U2 snRNP binding to the branch-point sequences of introns through interaction with the U2AF large subunit (U2AF65). However, the functions of U2AF35 in plants are poorly understood. In this study, we discovered that the OsU2AF35a gene was vigorously induced by heat stress and could positively regulate rice thermotolerance during both the seedling and reproductive growth stages. OsU2AF35a interacts with OsU2AF65a within the nucleus, and both of them can form condensates through liquid-liquid phase separation (LLPS) following heat stress. The intrinsically disordered regions (IDR) are accountable for their LLPS. OsU2AF35a condensation is indispensable for thermotolerance. RNA-seq analysis disclosed that, subsequent to heat treatment, the expression levels of several genes associated with water deficiency and oxidative stress in osu2af35a-1 were markedly lower than those in ZH11. In accordance with this, OsU2AF35a is capable of positively regulating the oxidative stress resistance of rice. The pre-mRNAs of a considerable number of genes in the osu2af35a-1 mutant exhibited defective splicing, among which was the OsHSA32 gene. Knocking out OsHSA32 significantly reduced the thermotolerance of rice, while overexpressing OsHSA32 could partially rescue the heat sensitivity of osu2af35a-1. Together, our findings uncovered the essential role of OsU2AF35a in rice heat stress response through protein separation and regulating alternative pre-mRNA splicing.

The mechanized direct seeding of rice (Oryza sativa L.) is a major trend nowadays. The elongation of rice mesocotyl and coleoptile can facilitate the rapid emergence of seedlings under deep mechanized sowing. Currently, most of the cultivated rice accessions have short mesocotyls or coleoptiles, with only a few related genes cloned. However, understanding and enhancing the ability of rice seedlings to rapidly emerge from deep sowing depths is crucial. Herein, we assessed 745 core rice germplasm accessions sown under a soil cover depth of 10 cm and found few long mesocotyl and coleoptile germplasms. We conducted genome-wide association study using six models to obtain three or more multi-model co-localization candidate regions and calculated Fst between the phenotypes of extreme samples to determine genetic differences. The candidate regions associated with mesocotyl and coleoptile lengths were identified by integrating Fst and multi-model localization results. This multi-model localization method may accelerate the mining of genes related to the mesocotyl and coleoptile, providing valuable targets for functional validation and marker-assisted selection in rice breeding programs. We utilized diverse GWAS models to co-locate candidate loci for mesocotyl and coleoptile elongation. We calculated Fst in phenotypically divergent materials to refine candidate regions. This multi-model localization method provides valuable targets for rice breeding and mechanized cultivation. Rice requires longer mesocotyl and coleoptile for rapid seedling emergence in direct seeding. It is necessary to identify key genes that control mesocotyl and coleoptile traits to facilitate the breeding of new varieties. We analyzed the mesocotyl and coleoptile lengths of 745 rice samples and conducted genome-wide association study using six different models to co-locate candidate loci. Several valuable candidate regions were consistently detected using different models. These regions included previously reported genes related to mesocotyl/coleoptile elongation and other agronomic traits such as plant height, heading date, and yield. Our findings validated the accuracy of the multi-model analysis and provided guidance on efficiently mining genes involved in mesocotyl/coleoptile elongation from those known genes controlling plant growth and development. This study may contribute to the breeding of new rice varieties suitable for direct-seeding by synergistic selection of multi-traits.

Given the increase in global climate change,reconciling the increasing demand for food production with dwindling water resources poses a formidable challenge.To ensure sustainable rice production and global food security,a thorough exploration of the genetic mechanism that underlies the delicate balance between growth and drought tolerance in rice is imperative.The sessile nat-ure of plants has driven their adaptive evolution,enabling them to regulate gene expression and exhibit physiological and develop-mental plasticity in response to environmental changes[1].Alter-native splicing(AS),a prevalent phenomenon in plants,plays a crucial role in the regulation of posttranscriptional gene expression[2].AS serves as an essential mechanism involved in both develop-mental processes and stress responses[3-5].

Soybean (Glycine max L.) is a protein and oil crop grown worldwide. Its fitness may be reduced by deleterious mutations, whose identification and purging is desirable for crop breeding. In the published whole-genome re-sequenced data of 2214 soybean accessions, including 221 wild soybean, 1132 landrace cultivars and 861 improved soybean lines, we identified 115,275 deleterious single-nucleotide polymorphisms (SNPs). Numbers of deleterious alleles increased from wild soybeans to landraces and decreased from landraces to modern improved lines. Genes in selective-sweep regions showed fewer deleterious mutations than the remaining genes. Deleterious mutations explained 4.3%–48% more phenotypic variation than randomly selected SNPs for resistance to soybean cyst nematode race 2 (SCN2), soybean cyst nematode race 3 (SCN3) and soybean mosaic virus race 3 (SMV3). These findings illustrate how mutation load has shifted during soybean domestication, expansion and improvement and provide candidate sites for breeding out deleterious mutations in soybean by genome editing and/or conventional breeding focused on the selection of progeny with fewer deleterious alleles.

To allay excessive public concern about the safety of transgenic foods, and to optimize insect-resistant genes expression to delay the evolution of resistance in pests, we developed a promising strategy to fuse the GOI (gene of interest) with OsrbcS (rice small subunit of ribulose bisphosphate carboxylase/oxygenase) in transgenic rice, which acted as a carrier, driven by the OsrbcS native promoter to sequester its expression in green tissues. Using eYFP as a trial, we reported a high-level accumulation of eYFP in green tissue and almost none in the seed and root of the fused construct compared to the non-fused construct. After applying this fusion strategy in insect-resistant rice breeding, recombinant OsrbcS-Cry1Ab/Cry1Ac expressed rice plants conferred high resistance to leaffolders and striped stem borers, among which two single-copy lines possessed normal agronomic performance in the field. Specifically, Cry1Ab/Cry1Ac protein levels in single-copy construct transgenic lines ranged from 1.8 to 11.5 µg g−1 in the leaf, higher than the Actin I promoter-driven control, T51-1, about 1.78 µg g−1 in the leaf, but negligible (only 0.00012–0.00117 µg g−1) in endosperm by ELISA analysis. Our study provided a novel approach to creating Cry1Ab/Cry1Ac-free endosperm rice with a high level of insect-resistant protein in green tissues through the simultaneous usage of the OsrbcS promoter and OsrbcS as a fusion partner.

Ubc13 is required for Lys63-linked polyubiquitination and innate immune responses in mammals, but its functions in plant immunity still remain largely unknown. Here, we used molecular biological, pathological, biochemical, and genetic approaches to evaluate the roles of rice OsUbc13 in response to pathogens. The OsUbc13-RNA interference (RNAi) lines with lesion mimic phenotypes displayed a significant increase in the accumulation of flg22- and chitin-induced reactive oxygen species, and in defence-related genes expression or hormones as well as resistance to Magnaporthe oryzae and Xanthomonas oryzae pv oryzae. Strikingly, OsUbc13 directly interacts with OsSnRK1a, which is the α catalytic subunit of SnRK1 (sucrose non-fermenting-1-related protein kinase-1) and acts as a positive regulator of broad-spectrum disease resistance in rice. In the OsUbc13-RNAi plants, although the protein level of OsSnRK1a did not change, its activity and ABA sensitivity were obviously enhanced, and the K63-linked polyubiquitination was weaker than that of wild-type Dongjin (DJ). Overexpression of the deubiquitinase-encoding gene OsOTUB1.1 produced similar effects with inhibition of OsUbc13 in affecting immunity responses, M. oryzae resistance, OsSnRK1a ubiquitination, and OsSnRK1a activity. Furthermore, re-interfering with OsSnRK1a in one OsUbc13-RNAi line (Ri-3) partially restored its M. oryzae resistance to a level between those of Ri-3 and DJ. Our data demonstrate OsUbc13 negatively regulates immunity against pathogens by enhancing the activity of OsSnRK1a.

Thousand grain weight (TGW) is an important determinant of rice yield, and correlates with grain size, plumpness and grain number per panicle. In rice, there are fewer association mapping studies relating grain weight traits using both SSR and SNP markers. In this study, in order to find robust SSR markers associated with TGW trait and mine elite accessions in rice, we investigated the TGW trait across six environments using a natural population consisted of 462 accessions, and then performed association mapping using both SSR and SNP markers. Using the six datasets from the six environments and their best linear unbiased estimator, we identified eight TGW associated SSR markers, with three environmentally stable and one newly found, on five chromosomes. The associated markers have genetic effect from 3.44% to 20.84%, and two of them carry stable elite allele with positive effect across different environments. Candidate interval association mapping using re-sequencing derived SNP/InDel markers further confirms the TGW-SSR association, and also suggests that 3 TGW-SSR associations were high confident in intervals of size from 176 to 603 kb. These results not only shed more lights on the genetics of TGW trait, but also suggest that the multi-allelic SSR markers should be used as an alternative power tool in gene or QTL mapping.

Mesocotyl elongation is a key trait influencing seedling emergence and establishment in direct-seeding rice cultivation. The phytohormone gibberellin (GA) has positive effects on mesocotyl elongation in rice. However, the physiological and molecular basis underlying the regulation of mesocotyl elongation mediated by GA priming under deep-sowing conditions remains largely unclear. In the present study, we performed a physiological and comprehensive transcriptomic analysis of the function of GA priming in mesocotyl elongation and seedling emergence using a direct-seeding japonica rice cultivar ZH10 at a 5-cm sowing depth. Physiological experiments indicated that GA priming significantly improved rice seedling emergence by increasing the activity of starch-metabolizing enzymes and compatible solute content to supply the energy essential for subsequent development. Transcriptomic analysis revealed 7074 differentially expressed genes (false discovery rate of <0.05, |log2(fold change)| of ⩾1) after GA priming. Furthermore, gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analyses revealed that genes associated with transcriptional regulation, plant hormone biosynthesis or signaling, and starch and sucrose metabolism were critical for GA-mediated promotion of rice mesocotyl elongation. Further analyses showed that the expression of the transcription factor (TF) genes (v-myb avian myeloblastosis viral oncogene homolog (MYB) alternative splicing 1 (MYBAS1), phytochrome-interacting factors 1 (PIF1), Oryza sativa teosinte branched 1/cycloidea/proliferating cell factor 5 (OsTCP5), slender 1 (SLN1), and mini zinc finger 1 (MIF1)), plant hormone biosynthesis or signaling genes (brassinazole-resistant 1 (BZR1), ent-kaurenoic acid oxidase-like (KAO), GRETCHEN HAGEN 3.2 (GH3.2), and small auxin up RNA 36 (SAUR36)), and starch and sucrose metabolism genes (α-amylases (AMY2A and AMY1.4)) was highly correlated with the mesocotyl elongation and deep-sowing tolerance response. These results enhance our understanding of how nutrient metabolism-related substances and genes regulate rice mesocotyl elongation. This may facilitate future studies on related genes and the development of novel rice varieties tolerant to deep sowing.

Stable and uniform expression of reference genes across samples plays a key role in accurate normalization of gene expression by reverse-transcription quantitative polymerase chain reaction (RT-qPCR). For rice study, there is still a lack of validation and recommendation of appropriate reference genes with high stability depending on experimental conditions. Eleven candidate reference genes potentially owning high stability were evaluated by geNorm and NormFinder for their expression stability in 22 various experimental conditions. Best combinations of multiple reference genes were recommended depending on experimental conditions, and the holistic stability of reference genes was also evaluated. Reference genes would become more variable and thus needed to be critically selected in experimental groups of tissues, heat, 6-benzylamino purine, and drought, but they were comparatively stable under cold, wound, and ultraviolet-B stresses. Triosephosphate isomerase (TI), profilin-2 (Profilin-2), ubiquitin-conjugating enzyme E2 (UBC), endothelial differentiation factor (Edf), and ADP-ribosylation factor (ARF) were stable in most of our experimental conditions. No universal reference gene showed good stability in all experimental conditions. To get accurate expression result, suitable combination of multiple reference genes for a specific experimental condition would be a better choice. This study provided an application guideline to select stable reference genes for rice gene expression study.

The ubiquitin (Ub)-conjugating enzyme, Ubc13, has been known to be involved in error-free DNA damage tolerance (or post-replication repair) via catalyzing Lys63-linked polyubiquitin chains formation together with a Ubc variant. However, its functions remain largely unknown in plant species, especially in monocotyledons. In this study, we cloned a Ub-conjugating enzyme, OsUbc13, that shares the conserved domain of Ubc with AtUBC13B in Oryza sativa L., which encodes a protein of 153 amino acids; the deduced sequence shares high similarities with other homologs. Real-time quantitative polymerase chain reaction (PCR) indicated that OsUbc13 transcripts could be detected in all tissues examined, and the expression level was higher in palea, pistil, stamen, and leaf, and lower in root, stem, and lemma; the expression of OsUbc13 was induced by low temperature, methylmethane sulfate (MMS), and H2O2, but repressed by mannitol, abscisic acid (ABA), and NaCl. OsUbc13 was probably localized in the plasma and nuclear membranes. About 20 proteins, which are responsible for the positive yeast two-hybrid interaction of OsUbc13, were identified. These include the confirmed OsVDAC (correlated with apoptosis), OsMADS1 (important for development of floral organs), OsB22EL8 (related to reactive oxygen species (ROS) scavenging and DNA protection), and OsCROC-1(required for formation of Lys63 polyubiquitylation and error-free DNA damage tolerance). The molecular characterizationprovides a foundation for the functional study of OsUbc13.

BTF3 has been recognized to be involved in plant growth and development. But its function remains mostly unknown during seed germination and seedling stage. Here, we have analyzed OsBTF3-related sequences inOryza sativaL. subspecies, japonica, which resembles with the conserved domain of a nascent polypeptide associated complex (NAC) with different homologs of OsBTF3 and human BTF3. Inhibition ofOsj10gBTF3has led to considerable morphological changes during seed germination and seedling growth. Germination percentage was not influenced by the application of GA3, ABA, and NaCl but all concentrations caused wild-type (WT) seeds to germinate more rapidly than the RNAi (Osj10gBTF3Ri) transgenic lines. Seedling inhibition was more severe in theOsj10gBTF3Riseedlings compared with their WT especially when treated with 100 or 200 μM GA3; 50% reduction in shoots was observed inOsj10gBTF3Riseedlings. The expression ofOsj3g1BTF3,Osj3g2BTF3andOsj10gBTF3was primarily constitutive and generally modulated by NaCl, ABA, and GA3stresses in bothOsj10gBTF3Rilines and WT at the early seedling stage, suggesting thatOsj3g1BTF3andOsj10gBTF3are much similar but different fromOsj3g2BTF3in biological function. These results show that OsBTF3 plays an important role in seed germination and seedling growth gives a new perception demonstrating that more multifaceted regulatory functions are linked with BTF3 in plants.

Protein ubiquitination is a fundamental post-translational modification event that serves as a signaling function in diverse biological processes.Ubiquitination is accomplished by a series of three enzymatic steps as follows : E1(ubiquitin-activating enzyme)-E2(ubiquitin-conjugating enzyme,Ubc)-E3(ubiquitin ligase enzyme).In these processes,the role of Ubc is of pivotal importance.All Ubcs have an active site cysteine(Cys) residue within the highly conserved UBC domain.Ubc-like proteins share high similarities with Ubc but lack the active site Cys which is essential for the conjugation and transfer of ubiquitin to protein substrates,therefore,they have also been designated as UEV(ubiquitin-conjugating E2enzyme variant) proteins.Ubc-like proteins have been found in all eukaryotes examined such as animal,plant and yeast.To date,the functions of Ubc-like protein have been extensively studied in animal and yeast.However,it remains largely unknown in plant species,especially in monocot plants,so it is necessary to study Ubc-like protein in this field.Rice is a prominent model for monocotyledonous plants and one of the most important food crops.However,no information is available on Ubc-like protein in rice or other monocot plants.Here,we mainly characterized the expression pattern and subcellular localization of a rice Ubc-like protein in order to provide a foundation for the function studies of UEV proteins in monocot plants.The molecular characterization of an Ubc-like protein gene in rice was cloned and analyzed,and was designated as OsCROC-1Ain terms of protein sequence analysis using bioinformatics,gene expression via real-time quantitative PCR,and fluorescence signal localization of the fusion protein in transformed tobacco suspension cultures.The open reading frame(ORF) of OsCROC-1A was cloned from rice(Oryza sativa,cultivar Nipponbare) via RT-PCR,and was used to analyze the expression pattern under abiotic stresses.OsCROC1Acontained five exons and four introns,and it encoded a UEV homolog of 146amino acids corresponding to a theoretical molecular mass of 17ku and pI of 6.42.OsCROC-1Aharbored a conserved UBC domain and D catalytic site,and the deduced sequence shared similarities with other homologs ranging from 42.61% to 84.14%.Real-time quantitative PCR was conducted to analyze the spatial expression pattern of OsCROC-1A as well as the changes of its expression level under abiotic stresses.As a result,OsCROC-1A was found to be expressed in all tissues examined,and the abundance level was high in leaf,root,lemma,and low in palea,stem,pistil,callus,and the lowest in stamen,implying that the OsCROC-1A is important to both vegetative growth and reproductive development in rice especially to that of leaf,root,lemma.The expression of OsCROC-1A was induced by low temperature of 4 ℃,20mmol / L H 2 O 2 and 0.01% MMS,whereas it was repressed by 300mmol / L mannitol,100 μ mol / L abscisic acid(ABA) and 200mmol / L NaCl,suggesting that OsCROC-1Acan respond actively to the negative circumstances by changing its transcripts.OsCROC-1A-EGFP was localized to both nucleus and cytosol,indicating that OsCROC1Anot only can play its role in the nucleus as a transcription factor,but also have function in cytosol.In conclusion,OsCROC-1A encodes an Ubc-like protein.It is constitutively expressed in all the tissues examined and is generally modulated by abiotic stress,and OsCROC-1Ahas functions in both nucleus and cytosol.These data provide a foundation for investigating the cellular function and molecular mechanism of OsCROC-1A.

BTF3, which was originally recognized as a basal transcription factor, has been known to be involved in transcription initiation, translational regulation and protein localization in many eukaryotic organisms. However, its function remains largely unknown in plant species. In the present study, we analyzed a BTF3-related sequence in Oryza sativa L. subsp. japonica, which shares the conserved domain of a nascent polypeptide-associated complex with human BTF3, and was referred to as Osj10gBTF3. The expression of Osj10gBTF3 was primarily constitutive and generally modulated by salt, high temperature and exogenous phytohormone stress. The Osj10gBTF3::EGFP (enhanced green fluorescence protein) fusion protein was localized in both the nucleus and cytoplasmic membrane system. Inhibition of Osj10gBTF3 led to significant morphological changes in all detected tissues and organs, with a reduced size of between 25% and 52%. Furthermore, the pollen that developed was completely sterile, which was correlated with the altered expression of two Rf (fertility restorer)-like genes that encode pentatricopeptide repeat-containing proteins OsPPR676 and OsPPR920, translational initiation factors OseIF3e and OseIF3h, and the heat shock protein OsHSP82. These findings were verified through a yeast two-hybrid assay using a Nipponbare callus cDNA library as bait followed by the reverse transcription-PCR analysis of total leaf or anther RNAs. Our demonstration of the important role of Osj10gBTF3 in rice growth and development provides new insights showing that more complex regulatory functions are associated with BTF3 in plants.

In an earlier greenhouse screening, we identified a local indica cultivar HT54 tolerant to high temperature at both seedling and grain-filling stages. In this study, we develop an optimized procedure for fine assessment of this heat tolerance. The results indicated that HT54 seedlings could tolerate high temperature up to 48 °C for 79h. The genetic analysis of F(1) and F(2) offspring derived from the cross between HT54 and HT13, a heat-sensitive breeding line, reveals that the heat tolerance of HT54 was controlled by a dominant major locus, which has been designated as OsHTAS (Oryza sativa heat tolerance at seedling stage). This locus was mapped on rice chromosome 9 within an interval of 420kb between markers of InDel5 and RM7364. The determined candidate ZFP gene has been confirmed to be cosegregated with a single nucleotide polymorphism (SNP) developed PCR-restriction fragment length polymorphism (RFLP) marker RBsp1407 in its promoter region. Another heat tolerance-associated SNP was identified in the first intron of its 5'-untranslated region. The existence of these SNPs thereby indicated that the OsHTAS locus contains at least two alleles. We named the one from HT54 as OsHTAS ( a ) and the one from HT13 as OsHTAS ( b ). Further dynamic expression analysis demonstrated that OsHTAS ( a ) was actively responsive to 45 °C high temperature stress compared with the OsHTAS ( b ) allele.

The MADS-box gene family encodes conserved transcription factors and functions not only in reproductive development, but also in vegetative growth. In this study, a rice MADS-box gene OsMADS15, an ortholog of Arabidopsis AP1 gene, has been functionally characterized. Rice seedlings overexpressing OsMADS15 showed precocious phenotypes of early internode elongation, shoot-borne crown root development, reduced plant height and early flowering. The axillary buds developments in OsMADS15 overexpressors were accelerated, and the buds frequently grew into effective tillers. The panicles of OsMADS15 transgenic rice plants were largely compromised for growth and branching in comparison with wild type. In the tillering stage, the OsMADS15 overexpression rice plants tillered later and less than the wild type, and in the maturity stage, the culms of the overexpression lines bore more stem nodes. Quantitative polymerase chain reaction (PCR) results showed that expression levels of WUSCHEL-related Homeobox (WOX) gene, WOX11, and some flowering regulators were promoted in the OsMADS15 overexpression transgenic plants, indicating that OsMADS15 had a wide range of regulations. These results clearly indicate that OsMADS15 plays important roles not only in transition to reproductive development, but also in crown root development.