Identification of Salt-Sensitive and Salt-Tolerant Genes through Weighted Gene Co-Expression Networks across Multiple Datasets: A Centralization and Differential Correlation Analysis.

Salt stress is a significant challenge that severely hampers rice growth, resulting in decreased yield and productivity. Over the years, researchers have identified biomarkers associated with salt stress to enhance rice tolerance. However, the understanding of the mechanism underlying salt tolerance in rice remains incomplete due to the involvement of multiple genes. Given the vast amount of genomics and transcriptomics data available today, it is crucial to integrate diverse datasets to identify key genes that play essential roles during salt stress in rice. In this study, we propose an integration of multiple datasets to identify potential key transcription factors. This involves utilizing network analysis based on weighted co-expression networks, focusing on gene-centric measurement and differential co-expression relationships among genes. Consequently, our analysis reveals 86 genes located in markers from previous meta-QTL analysis. Moreover, six transcription factors, namely LOC_Os03g45410 (OsTBP2), LOC_Os07g42400 (OsGATA23), LOC_Os01g13030 (OsIAA3), LOC_Os05g34050 (OsbZIP39), LOC_Os09g29930 (OsBIM1), and LOC_Os10g10990 (transcription initiation factor IIF), exhibited significantly altered co-expression relationships between salt-sensitive and salt-tolerant rice networks. These identified genes hold potential as crucial references for further investigation into the functions of salt stress response in rice plants and could be utilized in the development of salt-resistant rice cultivars. Overall, our findings shed light on the complex genetic regulation underlying salt tolerance in rice and contribute to the broader understanding of rice's response to salt stress.

Association of a Specific OsCULLIN3c Haplotype with Salt Stress Responses in Local Thai Rice.

We previously found that OsCUL3c is involved in the salt stress response. However, there are no definitive reports on the diversity of OsCUL3c in local Thai rice. In this study, we showed that the CUL3 group was clearly separated from the other CUL groups; next, we focused on OsCUL3c, the third CUL3 of the CUL3 family in rice, which is absent in Arabidopsis. A total of 111 SNPs and 28 indels over the OsCUL3c region, representing 79 haplotypes (haps), were found. Haplotyping revealed that group I (hap A and hap C) and group II (hap B1 and hap D) were different mutated variants, which showed their association with phenotypes under salt stress. These results were supported by cis-regulatory elements (CREs) and transcription factor binding sites (TFBSs) analyses. We found that LTR, MYC, [AP2; ERF], and NF-YB, which are related to salt stress, drought stress, and the response to abscisic acid (ABA), have distinct positions and numbers in the haplotypes of group I and group II. An RNA Seq analysis of the two predominant haplotypes from each group showed that the OsCUL3c expression of the group I representative was upregulated and that of group II was downregulated, which was confirmed by RT-qPCR. Promoter changes might affect the transcriptional responses to salt stress, leading to different regulatory mechanisms for the expression of different haplotypes. We speculate that OsCUL3c influences the regulation of salt-related responses, and haplotype variations play a role in this regulation.

OsBTBZ1 Confers Salt Stress Tolerance in Arabidopsis thaliana.

Rice (Oryza sativa L.), one of the most important commodities and a primary food source worldwide, can be affected by adverse environmental factors. The chromosome segment substitution line 16 (CSSL16) of rice is considered salt-tolerant. A comparison of the transcriptomic data of the CSSL16 line under normal and salt stress conditions revealed 511 differentially expressed sequence (DEseq) genes at the seedling stage, 520 DEseq genes in the secondary leaves, and 584 DEseq genes in the flag leaves at the booting stage. Four BTB genes, OsBTBZ1, OsBTBZ2, OsBTBN3, and OsBTBN7, were differentially expressed under salt stress. Interestingly, only OsBTBZ1 was differentially expressed at the seedling stage, whereas the other genes were differentially expressed at the booting stage. Based on the STRING database, OsBTBZ1 was more closely associated with other abiotic stress-related proteins than other BTB genes. The highest expression of OsBTBZ1 was observed in the sheaths of young leaves. The OsBTBZ1-GFP fusion protein was localized to the nucleus, supporting the hypothesis of a transcriptionally regulatory role for this protein. The bt3 Arabidopsis mutant line exhibited susceptibility to NaCl and abscisic acid (ABA) but not to mannitol. NaCl and ABA decreased the germination rate and growth of the mutant lines. Moreover, the ectopic expression of OsBTBZ1 rescued the phenotypes of the bt3 mutant line and enhanced the growth of wild-type Arabidopsis under stress conditions. These results suggest that OsBTBZ1 is a salt-tolerant gene functioning in ABA-dependent pathways.

Genome-Wide Association Study of Starch Properties in Local Thai Rice.

Rice (Oryza sativa L.) is the main source of energy for humans and a staple food of high cultural significance for much of the world's population. Rice with highly resistant starch (RS) is beneficial for health and can reduce the risk of disease, especially type II diabetes. The identification of loci affecting starch properties will facilitate breeding of high-quality and health-supportive rice. A genome-wide association study (GWAS) of 230 rice cultivars was used to identify candidate loci affecting starch properties. The apparent amylose content (AAC) among rice cultivars ranged from 7.04 to 33.06%, and the AAC was positively correlated with RS (R2 = 0.94) and negatively correlated with rapidly available glucose (RAG) (R2 = -0.73). Three loci responsible for starch properties were detected on chromosomes 1, 6, and 11. On chromosome 6, the most significant SNP corresponded to LOC_Os06g04200 which encodes granule-bound starch synthase I (GBSSI) or starch synthase. Two novel loci associated with starch traits were LOC_Os01g65810 and LOC_Os11g01580, which encode an unknown protein and a sodium/calcium exchanger, respectively. The markers associated with GBSSI and LOC_Os11g01580 were tested in two independent sets of rice populations to confirm their effect on starch properties. The identification of genes associated with starch traits will further the understanding of the molecular mechanisms affecting starch in rice and may be useful in the selection of rice varieties with improved starch.

Ectopic expression of rice malate synthase in Arabidopsis revealed its roles in salt stress responses.

Expression of rice malate synthase (OsMS), one of the two key genes in the glyoxylate cycle, is highly upregulated under salt stress. In this study, we aimed to investigate the role of OsMS in salt stress responses using the Arabidopsis T-DNA insertional mutant line of malate synthase (AtMS), an OsMS orthologous gene, for ectopic expression. Germination of the Atms mutant under salt stress was lower than that of Arabidopsis Col-0 wildtype (WT); furthermore, the two Atms mutant lines ectopically expressing OsMS reversed the salt-sensitive phenotype. Atms mutants salt-treated for 3 days exhibited higher electrolyte leakage, higher Na+/K+ ratio, lower expression of stress-responsive genes, and lower soluble sugar content than WT and the two OsMS-expressing Atms mutant lines. Consistently, Atms mutants salt-treated for 3 days followed by a 5-day recovery period displayed greater fresh-weight reduction. Notably, leaf greenness and chlorophyll and total carotenoid contents were higher in the Atms mutant lines than in the WT under stress. OsMS-expressing Atms mutants exhibited a change in pigment content closer to that of WT. During dark-induced senescence, an Atms mutant, Aticl, mutant (the other key gene in the glyoxylate cycle), and three double mutant lines of Atms and Aticl exhibited lower decreases in leaf greenness than the WT and OsMS-expressing Atms mutant lines. Furthermore, SAG12 expression levels in the Atms mutant, Aticl mutant, and three double mutant lines were lower than those in OsMS-expressing Atms mutant lines. Altogether, our data indicate that OsMS likely plays a key role in salt stress responses, possibly through the induction of leaf senescence.

Changes in physiological responses of OsCaM1-1 overexpression in the transgenic rice under dehydration stress.

Calmodulin, a primary calcium sensor in eukaryotes, binds calcium and regulates the activity of effector proteins in response to calcium signals that evoked in response to abiotic and biotic stress. To identify physiological responses associated with improved tolerance under dehydration stress that may be regulated by calmodulin in rice, the transgenic rice overexpressing OsCaM1-1, the control, and the wild-type KDML105 differing in their dehydration tolerance were compared 24 h after exposure to dehydration stress. The results demonstrated a greater increase in relative water content, relative growth rate, abscisic acid, photosynthetic pigment and proline contents, and antioxidant activities in the transgenic rice plants, whereas Na/K and Na/Ca ratio, lipid peroxidation, and electrolytic leakage decreased. The OsCaM1-1 gene overexpression in the transgenic rice showed greater tolerance to dehydration stress than non-transgenic rice, suggesting that OsCaM1-1 might play an important role in mitigating dehydration stress.

Hyperspectral and genome-wide association analyses of leaf phosphorus status in local Thai indica rice.

Phosphorus (P) is an essential mineral nutrient and one of the key factors determining crop productivity. P-deficient plants exhibit visual leaf symptoms, including chlorosis, and alter spectral reflectance properties. In this study, we evaluated leaf inorganic phosphate (Pi) contents, plant growth and reflectance spectra (420-790 nm) of 172 Thai rice landrace varieties grown hydroponically under three different P supplies (overly sufficient, mildly deficient and severely deficient conditions). We reported correlations between Pi contents and reflectance ratios computed from two wavebands in the range of near infrared (720-790 nm) and visible energy (green-yellow and red edge) (r > 0.69) in Pi-deficient leaves. Artificial neural network models were also developed which could classify P deficiency levels with 85.60% accuracy and predict Pi content with R2 of 0.53, as well as highlight important waveband sections. Using 217 reflectance ratio indices to perform genome-wide association study (GWAS) with 113,114 SNPs, we identified 11 loci associated with the spectral reflectance traits, some of which were also associated with the leaf Pi content trait. Hyperspectral measurement offers a promising non-destructive approach to predict plant P status and screen large germplasm for varieties with high P use efficiency.

Identification of a Negative Regulator for Salt Tolerance at Seedling Stage via a Genome-Wide Association Study of Thai Rice Populations.

Salt stress is a major limiting factor in crop production and yield in many regions of the world. The objective of this study was to identify the genes responsible for salt tolerance in Thai rice populations. We performed a genome-wide association study with growth traits, relative water content, and cell membrane stability at the seedling stage, and predicted 25 putative genes. Eleven of them were located within previously reported salt-tolerant QTLs (ST-QTLs). OsCRN, located outside the ST-QTLs, was selected for gene characterization using the Arabidopsis mutant line with T-DNA insertion in the orthologous gene. Mutations in the AtCRN gene led to the enhancement of salt tolerance by increasing the ability to maintain photosynthetic pigment content and relative water content, while the complemented lines with ectopic expression of OsCRN showed more susceptibility to salt stress detected by photosynthesis performance. Moreover, the salt-tolerant rice varieties showed lower expression of this gene than the susceptible rice varieties under salt stress conditions. The study concludes that by acting as a negative regulator, OsCRN plays an important role in salt tolerance in rice.

Salt stress responses and SNP-based phylogenetic analysis of Thai rice cultivars.

Genetic diversity is important for developing salt-tolerant rice (Oryza sativa L.) cultivars. Certain Thai rice accessions display salt tolerance at the adult or reproductive stage, but their response to salinity at the seedling stage is unknown. In this study, a total of 10 rice cultivars/line, including eight Thai cultivars and standard salt-tolerant cultivar and susceptible line, were screened using a hydroponic system to identify salt-tolerant genotypes at the seedling stage. Different morphophysiological indicators were used to classify tolerant and susceptible genotypes. Phylogenetic analyses were performed to determine the evolutionary relationships between the cultivars. Results showed that 'Lai Mahk', 'Jao Khao', 'Luang Pratahn', and 'Ma Gawk' exhibited salt stress tolerance comparable with the standard salt-tolerance check 'Pokkali'. Whole-exome single-nucleotide polymorphism (SNP)-based phylogenetic analysis showed that the Thai rice cultivars were monophyletic and distantly related to Pokkali and IR29. Lai Mahk and Luang Pratahn were found closely related when using the whole-exome SNPs for the analysis. This is also the case for the analysis of SNPs from 164 salt-tolerance genes and transcription regulatory genes. The tolerant cultivars shared the same haplotype for 16 genes. Overall, the findings of this study identified four salt-stress-tolerant Thai rice cultivars, which could be used in rice breeding programs for salinity tolerance.

Identification of Key Genes in 'Luang Pratahn', Thai Salt-Tolerant Rice, Based on Time-Course Data and Weighted Co-expression Networks.

Salinity is an important environmental factor causing a negative effect on rice production. To prevent salinity effects on rice yields, genetic diversity concerning salt tolerance must be evaluated. In this study, we investigated the salinity responses of rice (Oryza sativa) to determine the critical genes. The transcriptomes of 'Luang Pratahn' rice, a local Thai rice variety with high salt tolerance, were used as a model for analyzing and identifying the key genes responsible for salt-stress tolerance. Based on 3' Tag-Seq data from the time course of salt-stress treatment, weighted gene co-expression network analysis was used to identify key genes in gene modules. We obtained 1,386 significantly differentially expressed genes in eight modules. Among them, six modules indicated a significant correlation within 6, 12, or 48h after salt stress. Functional and pathway enrichment analysis was performed on the co-expressed genes of interesting modules to reveal which genes were mainly enriched within important functions for salt-stress responses. To identify the key genes in salt-stress responses, we considered the two-state co-expression networks, normal growth conditions, and salt stress to investigate which genes were less important in a normal situation but gained more impact under stress. We identified key genes for the response to biotic and abiotic stimuli and tolerance to salt stress. Thus, these novel genes may play important roles in salinity tolerance and serve as potential biomarkers to improve salt tolerance cultivars.

Comprehensive genome-wide analysis of calmodulin-binding transcription activator (CAMTA) in Durio zibethinus and identification of fruit ripening-associated DzCAMTAs.

BACKGROUND: Fruit ripening is an intricate developmental process driven by a highly coordinated action of complex hormonal networks. Ethylene is considered as the main phytohormone that regulates the ripening of climacteric fruits. Concomitantly, several ethylene-responsive transcription factors (TFs) are pivotal components of the regulatory network underlying fruit ripening. Calmodulin-binding transcription activator (CAMTA) is one such ethylene-induced TF implicated in various stress and plant developmental processes. RESULTS: Our comprehensive analysis of the CAMTA gene family in Durio zibethinus (durian, Dz) identified 10 CAMTAs with conserved domains. Phylogenetic analysis of DzCAMTAs, positioned DzCAMTA3 with its tomato ortholog that has already been validated for its role in the fruit ripening process through ethylene-mediated signaling. Furthermore, the transcriptome-wide analysis revealed DzCAMTA3 and DzCAMTA8 as the highest expressing durian CAMTA genes. These two DzCAMTAs possessed a distinct ripening-associated expression pattern during post-harvest ripening in Monthong, a durian cultivar native to Thailand. The expression profiling of DzCAMTA3 and DzCAMTA8 under natural ripening conditions and ethylene-induced/delayed ripening conditions substantiated their roles as ethylene-induced transcriptional activators of ripening. Similarly, auxin-suppressed expression of DzCAMTA3 and DzCAMTA8 confirmed their responsiveness to exogenous auxin treatment in a time-dependent manner. Accordingly, we propose that DzCAMTA3 and DzCAMTA8 synergistically crosstalk with ethylene during durian fruit ripening. In contrast, DzCAMTA3 and DzCAMTA8 antagonistically with auxin could affect the post-harvest ripening process in durian. Furthermore, DzCAMTA3 and DzCAMTA8 interacting genes contain significant CAMTA recognition motifs and regulated several pivotal fruit-ripening-associated pathways. CONCLUSION: Taken together, the present study contributes to an in-depth understanding of the structure and probable function of CAMTA genes in the post-harvest ripening of durian.

Combining Genome and Gene Co-expression Network Analyses for the Identification of Genes Potentially Regulating Salt Tolerance in Rice.

Salinity stress tolerance is a complex polygenic trait involving multi-molecular pathways. This study aims to demonstrate an effective transcriptomic approach for identifying genes regulating salt tolerance in rice. The chromosome segment substitution lines (CSSLs) of "Khao Dawk Mali 105 (KDML105)" rice containing various regions of DH212 between markers RM1003 and RM3362 displayed differential salt tolerance at the booting stage. CSSL16 and its nearly isogenic parent, KDML105, were used for transcriptome analysis. Differentially expressed genes in the leaves of seedlings, flag leaves, and second leaves of CSSL16 and KDML105 under normal and salt stress conditions were subjected to analyses based on gene co-expression network (GCN), on two-state co-expression with clustering coefficient (CC), and on weighted gene co-expression network (WGCN). GCN identified 57 genes, while 30 and 59 genes were identified using CC and WGCN, respectively. With the three methods, some of the identified genes overlapped, bringing the maximum number of predicted salt tolerance genes to 92. Among the 92 genes, nine genes, OsNodulin, OsBTBZ1, OsPSB28, OsERD, OsSub34, peroxidase precursor genes, and three expressed protein genes, displayed SNPs between CSSL16 and KDML105. The nine genes were differentially expressed in CSSL16 and KDML105 under normal and salt stress conditions. OsBTBZ1 and OsERD were identified by the three methods. These results suggest that the transcriptomic approach described here effectively identified the genes regulating salt tolerance in rice and support the identification of appropriate QTL for salt tolerance improvement.

Prediction of Human-Plasmodium vivax Protein Associations From Heterogeneous Network Structures Based on Machine-Learning Approach.

Malaria caused by Plasmodium vivax can lead to severe morbidity and death. In addition, resistance has been reported to existing drugs in treating this malaria. Therefore, the identification of new human proteins associated with malaria is urgently needed for the development of additional drugs. In this study, we established an analysis framework to predict human-P. vivax protein associations using network topological profiles from a heterogeneous network structure of human and P. vivax, machine-learning techniques and statistical analysis. Novel associations were predicted and ranked to determine the importance of human proteins associated with malaria. With the best-ranking score, 411 human proteins were identified as promising proteins. Their regulations and functions were statistically analyzed, which led to the identification of proteins involved in the regulation of membrane and vesicle formation, and proteasome complexes as potential targets for the treatment of P. vivax malaria. In conclusion, by integrating related data, our analysis was efficient in identifying potential targets providing an insight into human-parasite protein associations. Furthermore, generalizing this model could allow researchers to gain further insights into other diseases and enhance the field of biomedical science.

Ca2+/Calmodulin Complex Triggers CAMTA Transcriptional Machinery Under Stress in Plants: Signaling Cascade and Molecular Regulation.

Calcium (Ca2+) ion is a critical ubiquitous intracellular second messenger, acting as a lead currency for several distinct signal transduction pathways. Transient perturbations in free cytosolic Ca2+ ([Ca2+]cyt) concentrations are indispensable for the translation of signals into adaptive biological responses. The transient increase in [Ca2+]cyt levels is sensed by an array of Ca2+ sensor relay proteins such as calmodulin (CaM), eventually leading to conformational changes and activation of CaM. CaM, in a Ca2+-dependent manner, regulates several transcription factors (TFs) that are implicated in various molecular, physiological, and biochemical functions in cells. CAMTA (calmodulin-binding transcription activator) is one such member of the Ca2+-loaded CaM-dependent family of TFs. The present review focuses on Ca2+ as a second messenger, its interaction with CaM, and Ca2+/CaM-mediated CAMTA transcriptional regulation in plants. The review recapitulates the molecular and physiological functions of CAMTA in model plants and various crops, confirming its probable involvement in stress signaling pathways and overall plant development. Studying Ca2+/CaM-mediated CAMTA TF will help in answering key questions concerning signaling cascades and molecular regulation under stress conditions and plant growth, thus improving our knowledge for crop improvement.

Drought-Tolerance Gene Identification Using Genome Comparison and Co-Expression Network Analysis of Chromosome Substitution Lines in Rice.

Drought stress limits plant growth and productivity. It triggers many responses by inducing changes in plant morphology and physiology. KDML105 rice is a key rice variety in Thailand and is normally grown in the northeastern part of the country. The chromosome segment substitution lines (CSSLs) were developed by transferring putative drought tolerance loci (QTLs) on chromosome 1, 3, 4, 8, or 9 into the KDML105 rice genome. CSSL104 is a drought-tolerant line with higher net photosynthesis and leaf water potential than KDML105 rice. The analysis of CSSL104 gene regulation identified the loci associated with these traits via gene co-expression network analysis. Most of the predicted genes are involved in the photosynthesis process. These genes are also conserved in Arabidopsis thaliana. Seven genes encoding chloroplast proteins were selected for further analysis through characterization of Arabidopsis tagged mutants. The response of these mutants to drought stress was analyzed daily for seven days after treatment by scoring green tissue areas via the PlantScreen™ XYZ system. Mutation of these genes affected green areas of the plant and stability index under drought stress, suggesting their involvement in drought tolerance.

Isocitrate lyase plays important roles in plant salt tolerance.

BACKGROUND: Isocitrate lyase (ICL) is a key enzyme in the glyoxylate cycle. In a previous study in rice, the expression of the ICL-encoding gene (OsICL) was highly induced by salt stress and its expression was enhanced in transgenic rice lines overexpressing OsCam1-1, a calmodulin (CaM)-encoding gene. CaM has been implicated in salt tolerance mechanisms in plants; however, the cellular mechanisms mediated by CaM are not clearly understood. In this study, the role of OsICL in plant salt tolerance mechanisms and the possible involvement of CaM were investigated using transgenic plants expressing OsICL or OsCam1-1. RESULTS: OsICL was highly expressed in senesced leaf and significantly induced by salt stress in three OsCam1-1 overexpressing transgenic rice lines as well as in wild type (WT). In WT young leaf, although OsICL expression was not affected by salt stress, all three transgenic lines exhibited highly induced expression levels. In Arabidopsis, salt stress had negative effects on germination and seedling growth of the AtICL knockout mutant (Aticl mutant). To examine the roles of OsICL we generated the following transgenic Arabidopsis lines: the Aticl mutant expressing OsICL driven by the native AtICL promoter, the Aticl mutant overexpressing OsICL driven by the 35SCaMV promoter, and WT overexpressing OsICL driven by the 35SCaMV promoter. Under salt stress, the germination rate and seedling fresh and dry weights of the OsICL-expressing lines were higher than those of the Aticl mutant, and the two lines with the icl mutant background were similar to the WT. The Fv/Fm and temperature of rosette leaves in the OsICL-expressing lines were less affected by salt stress than they were in the Aticl mutant. Finally, glucose and fructose contents of the Aticl mutant under salt stress were highest, whereas those of OsICL-expressing lines were similar to or lower than those of the WT. CONCLUSIONS: OsICL, a salt-responsive gene, was characterized in the transgenic Arabidopsis lines, revealing that OsICL expression could revert the salt sensitivity phenotypes of the Aticl knockout mutant. This work provides novel evidence that supports the role of ICL in plant salt tolerance through the glyoxylate cycle and the possible involvement of OsCam1-1 in regulating its transcription.

Comparison between the Transcriptomes of 'KDML105' Rice and a Salt-Tolerant Chromosome Segment Substitution Line.

'KDML105' rice, known as jasmine rice, is grown in northeast Thailand. The soil there has high salinity, which leads to low productivity. Chromosome substitution lines (CSSLs) with the 'KDML105' rice genetic background were evaluated for salt tolerance. CSSL18 showed the highest salt tolerance among the four lines tested. Based on a comparison between the CSSL18 and 'KDML105' transcriptomes, more than 27,000 genes were mapped onto the rice genome. Gene ontology enrichment of the significantly differentially expressed genes (DEGs) revealed that different mechanisms were involved in the salt stress responses between these lines. Biological process and molecular function enrichment analysis of the DEGs from both lines revealed differences in the two-component signal transduction system, involving LOC_Os04g23890, which encodes phototropin 2 (PHOT2), and LOC_Os07g44330, which encodes pyruvate dehydrogenase kinase (PDK), the enzyme that inhibits pyruvate dehydrogenase in respiration. OsPHOT2 expression was maintained in CSSL18 under salt stress, whereas it was significantly decreased in 'KDML105', suggesting OsPHOT2 signaling may be involved in salt tolerance in CSSL18. PDK expression was induced only in 'KDML105'. These results suggested respiration was more inhibited in 'KDML105' than in CSSL18, and this may contribute to the higher salt susceptibility of 'KDML105' rice. Moreover, the DEGs between 'KDML105' and CSSL18 revealed the enrichment in transcription factors and signaling proteins located on salt-tolerant quantitative trait loci (QTLs) on chromosome 1. Two of them, OsIRO2 and OsMSR2, showed the potential to be involved in salt stress response, especially, OsMSR2, whose orthologous genes in Arabidopsis had the potential role in photosynthesis adaptation under salt stress.

Comparative Genomic Analysis of Rice with Contrasting Photosynthesis and Grain Production under Salt Stress.

Unfavourable environmental conditions, including soil salinity, lead to decreased rice (Oryza sativa L.) productivity, especially at the reproductive stage. In this study, we examined 30 rice varieties, which revealed significant differences in the photosynthetic performance responses under salt stress conditions during the reproductive stage, which ultimately affected yield components after recovery. In rice with a correlation between net photosynthetic rate (PN) and intercellular CO2 concentration (Ci) under salt stress, PN was found to be negatively correlated with filled grain number after recovery. Applying stringent criteria, we identified 130,317 SNPs and 15,396 InDels between two "high-yield rice" varieties and two "low-yield rice" varieties with contrasting photosynthesis and grain yield characteristics. A total of 2,089 genes containing high- and moderate-impact SNPs or InDels were evaluated by gene ontology (GO) enrichment analysis, resulting in over-represented terms in the apoptotic process and kinase activity. Among these genes, 262 were highly expressed in reproductive tissues, and most were annotated as receptor-like protein kinases. These findings highlight the importance of variations in signaling components in the genome and these loci can serve as potential genes in rice breeding to produce a variety with salt avoidance that leads to increased yield in saline soil.

Genome-wide association study for salinity tolerance at the flowering stage in a panel of rice accessions from Thailand.

BACKGROUND: Salt stress, a major plant environmental stress, is a critical constraint for rice productivity. Dissecting the genetic loci controlling salt tolerance in rice for improving productivity, especially at the flowering stage, remains challenging. Here, we conducted a genome-wide association study (GWAS) of salt tolerance based on exome sequencing of the Thai rice accessions. RESULTS: Photosynthetic parameters and cell membrane stability under salt stress at the flowering stage; and yield-related traits of 104 Thai rice (Oryza sativa L.) accessions belonging to the indica subspecies were evaluated. The rice accessions were subjected to exome sequencing, resulting in 112,565 single nucleotide polymorphisms (SNPs) called with a minor allele frequency of at least 5%. LD decay analysis of the panel indicates that the average LD for SNPs at 20 kb distance from each other was 0.34 (r2), which decayed to its half value (~ 0.17) at around 80 kb. By GWAS performed using mixed linear model, two hundred loci containing 448 SNPs on exons were identified based on the salt susceptibility index of the net photosynthetic rate at day 6 after salt stress; and the number of panicles, filled grains and unfilled grains per plant. One hundred and forty six genes, which accounted for 73% of the identified loci, co-localized with the previously reported salt quantitative trait loci (QTLs). The top four regions that contained a high number of significant SNPs were found on chromosome 8, 12, 1 and 2. While many are novel, their annotation is consistent with potential involvement in plant salt tolerance and in related agronomic traits. These significant SNPs greatly help narrow down the region within these QTLs where the likely underlying candidate genes can be identified. CONCLUSIONS: Insight into the contribution of potential genes controlling salt tolerance from this GWAS provides further understanding of salt tolerance mechanisms of rice at the flowering stage, which can help improve yield productivity under salinity via gene cloning and genomic selection.

Downstream components of the calmodulin signaling pathway in the rice salt stress response revealed by transcriptome profiling and target identification.

BACKGROUND: Calmodulin (CaM) is an important calcium sensor protein that transduces Ca2+ signals in plant stress signaling pathways. A previous study has revealed that transgenic rice over-expressing the calmodulin gene OsCam1-1 (LOC_Os03g20370) is more tolerant to salt stress than wild type. To elucidate the role of OsCam1-1 in the salt stress response mechanism, downstream components of the OsCam1-1-mediated response were identified and investigated by transcriptome profiling and target identification. RESULTS: Transcriptome profiling of transgenic 'Khao Dawk Mali 105' rice over-expressing OsCam1-1 and wild type rice showed that overexpression of OsCam1-1 widely affected the expression of genes involved in several cellular processes under salt stress, including signaling, hormone-mediated regulation, transcription, lipid metabolism, carbohydrate metabolism, secondary metabolism, photosynthesis, glycolysis, tricarboxylic acid (TCA) cycle and glyoxylate cycle. Under salt stress, the photosynthesis rate in the transgenic rice was slightly lower than in wild type, while sucrose and starch contents were higher, suggesting that energy and carbon metabolism were affected by OsCam1-1 overexpression. Additionally, four known and six novel CaM-interacting proteins were identified by cDNA expression library screening with the recombinant OsCaM1. GO terms enriched in their associated proteins that matched those of the differentially expressed genes affected by OsCam1-1 overexpression revealed various downstream cellular processes that could potentially be regulated by OsCaM1 through their actions. CONCLUSIONS: The diverse cellular processes affected by OsCam1-1 overexpression and possessed by the identified CaM1-interacting proteins corroborate the notion that CaM signal transduction pathways compose a complex network of downstream components involved in several cellular processes. These findings suggest that under salt stress, CaM activity elevates metabolic enzymes involved in central energy pathways, which promote or at least maintain the production of energy under the limitation of photosynthesis.