The QTL and Candidate Genes Regulating the Early Tillering Vigor Traits of Late-Season Rice in Double-Cropping Systems.

Rice effective panicle is a major trait for grain yield and is affected by both the genetic tiller numbers and the early tillering vigor (ETV) traits to survive environmental adversities. The mechanism behind tiller bud formation has been well described, while the genes and the molecular mechanism underlying rice-regulating ETV traits are unclear. In this study, the candidate genes in regulating ETV traits have been sought by quantitative trait locus (QTL) mapping and bulk-segregation analysis by resequencing method (BSA-seq) conjoint analysis using rice backcross inbred line (BIL) populations, which were cultivated as late-season rice of double-cropping rice systems. By QTL mapping, seven QTLs were detected on chromosomes 1, 3, 4, and 9, with the logarithm of the odds (LOD) values ranging from 3.52 to 7.57 and explained 3.23% to 12.98% of the observed phenotypic variance. By BSA-seq analysis, seven QTLs on chromosomes 1, 2, 4, 5, 7, and 9 were identified using single-nucleotide polymorphism (SNP) and insertions/deletions (InDel) index algorithm and Euclidean distance (ED) algorithm. The overlapping QTL resulting from QTL mapping and BSA-seq analysis was shown in a 1.39 Mb interval on chromosome 4. In the overlap interval, six genes, including the functional unknown genes Os04g0455650, Os04g0470901, Os04g0500600, and ethylene-insensitive 3 (Os04g0456900), sialyltransferase family domain containing protein (Os04g0506800), and ATOZI1 (Os04g0497300), showed the differential expression between ETV rice lines and late tillering vigor (LTV) rice lines and have a missense base mutation in the genomic DNA sequences of the parents. We speculate that the six genes are the candidate genes regulating the ETV trait in rice, which provides a research basis for revealing the molecular mechanism behind the ETV traits in rice.

Cytosine methylations in the promoter regions of genes involved in the cellular oxidation equilibrium pathways affect rice heat tolerance.

BACKGROUND: High temperatures, particularly at night, decrease rice yield and quality. As high nighttime temperatures (HNTs) become increasingly frequent due to climate change, it is imperative to develop rice crops that tolerate HNTs. DNA methylation may represent a potential avenue for HNT-tolerant rice strain development, as this mechanism regulates gene activity and cellular phenotype in response to adverse environmental conditions without changing the nucleotide sequence. RESULTS: After HNT exposure, the methylation patterns of cytosines in the CHH context differed noticeably between two coisogenic rice strains with significantly different levels in heat tolerance. Methylation differences between strains were primarily observed on successive cytosines in the promoter or downstream regions of transcription factors and transposon elements. In contrast to the heat-sensitive rice strain, the regions 358-359 bp and 2-60 bp downstream of two basal transcriptional factors (TFIID subunit 11 and mediator of RNA polymerase II transcription subunit 31, respectively) were fully demethylated in the heat-tolerant strain after HNT exposure. In the heat-tolerant strain, HNTs reversed the methylation patterns of successive cytosines in the promoter regions of various genes involved in abscisic acid (ABA)-related reactive oxygen species (ROS) equilibrium pathways, including the pentatricopeptide repeat domain gene PPR (LOC_Os07g28900) and the homeobox domain gene homeobox (LOC_Os01g19694). Indeed, PRR expression was inhibited in heat-sensitive rice strains, and the methylation rates of the cytosines in the promoter region of PRR were greater in heat-sensitive strains as compared to heat-tolerant strains. CONCLUSIONS: After HNT exposure, cytosines in the CHH context were more likely than cytosines in other contexts to be methylated differently between the heat-sensitive and heat-tolerant rice strains. Methylation in the promoter regions of the genes associated with ABA-related oxidation and ROS scavenging improved heat tolerance in rice. Our results help to clarify the molecular mechanisms underlying rice heat tolerance.

High Nighttime Temperature Induces Antioxidant Molecule Perturbations in Heat-Sensitive and Heat-Tolerant Coisogenic Rice ( Oryza sativa) Strains.

Global warming-associated increases in temperature, particularly at nighttime, are detrimental to rice yield and quality. Metabolomic profiling was used to examine and compare the short-term extreme high nighttime temperature-induced molecular perturbations in rice ( Oryza sativa) coisogenic strains with contrasting heat-tolerances at the first stage of seed ripening. Compared to the heat-sensitive strain, antioxidant molecules were higher in abundance in the heat-tolerant strain, whereas the abundances of molecules involved in photosynthesis, nucleotide catabolism, and the S-adenosylmethionine (SAM) cycle varied only slightly. Thus, we proposed that the high abundance of antioxidant molecules in the heat-tolerant strain alleviated cellular oxidative stress, which protected photosynthesis, nucleotide catabolism, and the SAM cycle, leading to good grain filling.

Quantitative iTRAQ-based proteomic analysis of rice grains to assess high night temperature stress.

Rice yield and quality are adversely affected by increasing global surface temperature, and are strongly attributed to high night temperature (HNT) than high daytime temperature. However, the molecular mechanism underlying the heat-tolerant characteristics of rice remains unclear. In the present study, we compared the proteomes of heat-tolerant and -sensitive lines of rice at early milky stage using an iTRAQ method. We have identified 38 differentially expressed proteins between the two lines, of which 32 proteins have been functionally annotated in NCBI and/or the UniProt database. These proteins were then classified into seven functional subgroups, which include signal transduction, transcript regulation, oxidation, defense response, transport, energy metabolism, and biosynthesis. Further analysis indicated that HNT stress could disrupt the redox equilibrium of plant cells, which in turn triggers the calcium-dependent protein kinase and COP9 signalosome, thereby regulating downstream genes/proteins that are involved in the HNT response. The candidate proteins may provide genetic resources for the improvement of heat-tolerant characteristics in rice, and the proposed model for signal transduction and transcriptional regulation may facilitate in the elucidation of the molecular mechanism underlying the response to HNT stress in rice.

Transcriptome changes in rice (Oryza sativa L.) in response to high night temperature stress at the early milky stage.

BACKGROUND: Rice yield and quality are adversely affected by high temperatures, especially at night; high nighttime temperatures are more harmful to grain weight than high daytime temperatures. Unfortunately, global temperatures are consistently increasing at an alarming rate and the minimum nighttime temperature has increased three times as much as the corresponding maximum daytime temperature over the past few decades. RESULTS: We analyzed the transcriptome profiles for rice grain from heat-tolerant and -sensitive lines in response to high night temperatures at the early milky stage using the Illumina Sequencing method. The analysis results for the sequencing data indicated that 35 transcripts showed different expressions between heat-tolerant and -sensitive rice, and RT-qPCR analyses confirmed the expression patterns of selected transcripts. Functional analysis of the differentially expressed transcripts indicated that 21 genes have functional annotation and their functions are mainly involved in oxidation-reduction (6 genes), metabolic (7 genes), transport (4 genes), transcript regulation (2 genes), defense response (1 gene) and photosynthetic (1 gene) processes. Based on the functional annotation of the differentially expressed genes, the possible process that regulates these differentially expressed transcripts in rice grain responding to high night temperature stress at the early milky stage was further analyzed. This analysis indicated that high night temperature stress disrupts electron transport in the mitochondria, which leads to changes in the concentration of hydrogen ions in the mitochondrial and cellular matrix and influences the activity of enzymes involved in TCA and its secondary metabolism in plant cells. CONCLUSIONS: Using Illumina sequencing technology, the differences between the transcriptomes of heat-tolerant and -sensitive rice lines in response to high night temperature stress at the early milky stage was described here for the first time. The candidate transcripts may provide genetic resources that may be useful in the improvement of heat-tolerant characters of rice. The model proposed here is based on differences in expression and transcription between two rice lines. In addition, the model may support future studies on the molecular mechanisms underlying plant responses to high night temperatures.

Comparative proteomic analysis of differentially expressed proteins in the early milky stage of rice grains during high temperature stress.

Rice yield and quality are adversely affected by high temperatures, and these effects are more pronounced at the 'milky stage' of the rice grain ripening phase. Identifying the functional proteins involved in the response of rice to high temperature stress may provide the basis for improving heat tolerance in rice. In the present study, a comparative proteomic analysis of paired, genetically similar heat-tolerant and heat-sensitive rice lines was conducted. Two-dimensional electrophoresis (2-DE) revealed a total of 27 differentially expressed proteins in rice grains, predominantly from the heat-tolerant lines. The protein profiles clearly indicated variations in protein expression between the heat-tolerant and heat-sensitive rice lines. Matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF MS) analysis revealed that 25 of the 27 differentially displayed proteins were homologous to known functional proteins. These homologous proteins were involved in biosynthesis, energy metabolism, oxidation, heat shock metabolism, and the regulation of transcription. Seventeen of the 25 genes encoding the differentially displayed proteins were mapped to rice chromosomes according to the co-segregating conditions between the simple sequence repeat (SSR) markers and the target genes in recombinant inbred lines (RILs). The proteins identified in the present study provide a basis to elucidate further the molecular mechanisms underlying the adaptation of rice to high temperature stress.

Identification of candidate genes related to rice grain weight under high-temperature stress.

The rise of global warming presents a problem for all living organisms, including rice and other staple plants. High temperatures impair rice grain weight by inhibiting the filling of the caryopses during the milky stage. The molecular mechanism behind this process, however, is poorly understood. Identifying candidate genes involved in responses to high-temperature stress may provide a basis for the improvement of heat tolerance in rice. Using paired, genetically similar heat-tolerant and heat-sensitive rice lines as plant materials, cDNA-AFLP analysis revealed a total of 54 transcript derived fragments (TDFs), mainly from the heat-tolerant lines. This clearly indicated variations in gene expression between the two rice lines. BLAST results showed that 28 of the 54 TDFs were homologous sequences. These homologous genes were found to encode proteins involved in signal transduction, oxidation, transcriptional regulation, transport, and metabolism. The functions and differential expression patterns of some important genes are further discussed. High temperature stress may trigger a wide range of changes in gene expression in rice caryopses, in turn affecting functions ranging from signal transduction to cellular metabolism. Forty-five of the 54 TDFs were mapped to rice chromosomes. The genes identified in the present study would make good candidates for further study into the molecular mechanisms underlying rice adaptation to high-temperature stress.

Evaluation of protocols used in 2-d electrophoresis for proteome analysis of young rice caryopsis.

In order to obtain a high-resolution electrophorogram of rice young panicle proteome, we evaluated various protocols commonly used in two-dimensional (2D) polyacrylamide gel electrophoresis (PAGE) of proteins, including gel staining protocol, pH range of immobilized pH gradient (IPG) strips and sample loading quantity. Results showed that a silver staining protocol using sensitized solution containing glacial acetic acid, sodium acetate and sodium thiosulfate (reported by Heukeshoven and Dernick in 1988) and a Coomassie Brilliant Blue staining method using solution containing G-250, ammonium sulfate and phosphoric acid (reported by Pink et al in 2010) demonstrated the superior staining effect. In addition, we also showed that higher resolution was achieved when IPG gel strip with pH range of 5-8 was used, compared to that with pH range of 4-7. Finally, the optimal loading quantity was determined as 130 µg using the 17 cm-long nonlinear IPG strip with pH 5-8 in combination with the silver nitrate staining protocol. The evaluated results would be helpful in proteome analysis of young rice caryopsis.