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Identification of GCC-box and TCC-box motifs in the promoters of differentially expressed genes in rice (Oryza sativa L.): Experimental and computational approaches.

ABSTRACT: The transcription factor selectively binds with the cis-regulatory elements of the promoter and regulates the differential expression of genes. In this study, we aimed to identify and validate the presence of GCC-box and TCC-box motifs in the promoters of upregulated differentially expressed genes (UR-DEGs) and downregulated differentially expressed genes (DR-DEGs) under anoxia using molecular beacon probe (MBP) based real-time PCR. The GCC-box motif was detected in UR-DEGs (DnaJ and 60S ribosomal protein L7 genes), whereas, the TCC-box was detected in DR-DEGs (DnaK and CPuORF11 genes). In addition, the mechanism of interaction of AP2/EREBP family transcription factor (LOC_Os03g22170) with GCC-box promoter motif present in DnaJ gene (LOC_Os06g09560) and 60S ribosomal protein L7 gene (LOC_Os08g42920); and TCC-box promoter motif of DnaK gene (LOC_Os02g48110) and CPuORF11 gene (LOC_Os02g01240) were explored using molecular dynamics (MD) simulations analysis including binding free energy calculations, principal component analyses, and free energy landscapes. The binding free energy analysis revealed that AP2/EREBP model residues such as Arg68, Arg72, Arg83, Lys87, and Arg90 were commonly involved in the formation of hydrogen bonds with GCC and TCC-box promoter motifs, suggesting that these residues are critical for strong interaction. The movement of the entire protein bound to DNA was restricted, confirming the stability of the complex. This study provides comprehensive binding information and a more detailed view of the dynamic interaction between proteins and DNA.

SUBMITTER: Prajapati GK

PROVIDER: S-EPMC6485614 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Identification of GCC-box and TCC-box motifs in the promoters of differentially expressed genes in rice (Oryza sativa L.): Experimental and computational approaches.

Prajapati Gopal Kumar GK Pandey Bharati B Mishra Awdhesh Kumar AK Baek Kwang-Hyun KH Pandey Dev Mani DM

PloS one 20190426 4

The transcription factor selectively binds with the cis-regulatory elements of the promoter and regulates the differential expression of genes. In this study, we aimed to identify and validate the presence of GCC-box and TCC-box motifs in the promoters of upregulated differentially expressed genes (UR-DEGs) and downregulated differentially expressed genes (DR-DEGs) under anoxia using molecular beacon probe (MBP) based real-time PCR. The GCC-box motif was detected in UR-DEGs (DnaJ and 60S ribosom ...[more]

PMID: 31026257

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Project description:Rice (Oryza sativa) is one of the most important cereal crops, providing food for more than half of the world's population. However, grain yields are challenged by various abiotic stresses such as drought, fertilizer, heat, and their interaction. Rice at reproductive stage is much more sensitive to environmental temperatures, and little is known about molecular mechanisms of rice spikelet in response to high temperature interacting with nitrogen (N).Here we reported the transcriptional profiling analysis of rice spikelet at meiosis stage using RNA sequencing (RNA-seq) as an attempt to gain insights into molecular events associated with temperature and nitrogen. This study received four treatments: 1) NN: normal nitrogen level (165 kg ha(-1)) with natural temperature (30 °C); 2) HH: high nitrogen level (330 kg ha(-1)) with high temperature (37 °C); 3) NH: normal nitrogen level and high temperature; and 4) HN: high nitrogen level and natural temperature, respectively. The de novo assembly generated 52,553,536 clean reads aligned with 72,667 unigenes. About 10 M reads were identified from each treatment. In these differentially expressed genes (DEGs), we found 151 and 323 temperature-responsive DEGs in NN-vs-NH and HN-vs-HH, and 114 DEGs were co-expressed. Meanwhile, 203 and 144 nitrogen-responsive DEGs were focused in NN-vs-HN and NH-vs-HH, and 111 DEGs were co-expressed. The temperature-responsive genes were principally associated with calcium-dependent protein, cytochrome, flavonoid, heat shock protein, peroxidase, ubiquitin, and transcription factor while the nitrogen-responsive genes were mainly involved in glutamine synthetase, transcription factor, anthocyanin, amino acid transporter, leucine zipper protein, and hormone. It is noted that, rice spikelet fertility was significantly decreased under high temperature, but it was more reduced under higher nitrogen. Accordingly, numerous spikelet genes involved in pollen development, pollen tube growth, pollen germination, especially sporopollenin biosynthetic process, and pollen exine formation were mainly down-regulated under high temperature. Moreover, the expression levels of co-expressed DEGs including 5 sporopollenin biosynthetic process and 7 pollen exine formation genes of NN-vs-NH were lower than that of HN-vs-HH. Therefore, these spikelet genes may play important roles in response to high temperature with high nitrogen and may be good candidates for crop improvement.This RNA-seq study will help elucidate the molecular mechanisms of rice spikelet defense response to high temperature interacting with high nitrogen level.

Project description:BackgroundRice (Oryza sativa L.), one of the most important food crop in the world, is considered to be a salt-sensitive crop. Excess levels of salt adversely affect all the major metabolic activities, including cell wall damage, cytoplasmic lysis and genomic stability. In order to cope with salt stress, plants have evolved high degrees of developmental plasticity, including adaptation via cascades of molecular networks and changes in gene expression profiles. Posttranscriptional regulation, through the activity of microRNAs, also plays an important role in the plant response to salinity conditions. MicroRNAs are small endogenous RNAs that modulate gene expression and are involved in the most essential physiological processes, including plant development and adaptation to environmental changes.ResultsIn the present study, we investigated the expression profiles of osa-MIR414, osa-MIR408 and osa-MIR164e along with their targeted genes, under salinity stress conditions in wild type and transgenic rice plants ectopically expressing the PDH45 (Pea DNA Helicase) gene. The present miRNAs were predicted to target the OsABP (ATP-Binding Protein), OsDSHCT (DOB1/SK12/helY-like DEAD-box Helicase) and OsDBH (DEAD-Box Helicase) genes, included in the DEAD-box helicase family. An in silico characterization of the proteins was performed and the miRNAs predicted targets were validated by RLM-5'RACE. The qRT-PCR analysis showed that the OsABP, OsDBH and OsDSHCT genes were up-regulated in response to 100 and 200 mM NaCl treatments. The present study also highlighted an increased accumulation of the gene transcripts in wild type plants, with the exception of the OsABP mRNA which showed the highest level (15.1-fold change compared to control) in the transgenic plants treated with 200 mM NaCl. Salinity treatments also affected the expression of osa-MIR414, osa-MIR164e and osa-MIR408, found to be significantly down-regulated, although the changes in miRNA expression were limited.ConclusionsOsa-MIR414, osa-MIR164e and osa-MIR408 were experimentally validated for the first time in plants as targeting the OsABP, OsDBH and OsDSHCT genes. Our data showed that that the genes were up-regulated and the miRNAs were down-regulated in relation to salt stress. The negative correlation between the miRNAs and their targets was proven.

Project description:BackgroundDirect seeding is an efficient cultivation technique in rice. However, poor low temperature germinability (LTG) of modern rice cultivars limits its application. Identifying the genes associated with LTG and performing molecular breeding is the fundamental way to address this issue. However, few LTG QTLs have been fine mapped and cloned so far.ResultsIn the present study, the LTG evaluation of 375 rice accessions selected from the Rice Diversity Panel 2 showed that there were large LTG variations within the population, and the LTG of Indica group was significantly higher than that of Japonica and Aus groups (p < 0.01). In total, eleven QTLs for LTG were identified through genome-wide association study (GWAS). Among them, qLTG_sRDP2-3/qLTG_JAP-3, qLTG_AUS-3 and qLTG_sRDP2-12 are first reported in the present study. The QTL on chromosome 10, qLTG_sRDP2-10a had the largest contribution to LTG variations in 375 rice accessions, and was further validated using single segment substitution line (SSSL). The presence of qLTG_sRDP2-10a could result in 59.8% increase in LTG under 15 °C low temperature. The expression analysis of the genes within qLTG_sRDP2-10a region indicated that LOC_Os10g22520 and LOC_Os10g22484 exhibited differential expression between the high and low LTG lines. Further sequence comparisons revealed that there were insertion and deletion sequence differences in the promoter and intron region of LOC_Os10g22520, and an about 6 kb variation at the 3' end of LOC_Os10g22484 between the high and low LTG lines, suggesting that the sequence variations of the two genes could be the cause for their differential expression in high and low LTG lines.ConclusionAmong the 11 QTLs identified in this study, qLTG_sRDP2-10a could also be detected in other three studies using different germplasm under different cold environments. Its large effect and stable expression make qLTG_sRDP2-10a particularly valuable in rice breeding. The two genes, LOC_Os10g22484 and LOC_Os10g22520, were considered as the candidate genes underlying qLTG_sRDP2-10a. Our results suggest that integrating GWAS and SSSL can facilitate identification of QTL for complex traits in rice. The identification of qLTG_sRDP2-10a and its candidate genes provide a promising source for gene cloning of LTG and molecular breeding for LTG in rice.

Project description:affy_meloidogyne_rice - affy_meloidogyne_rice - Plant-parasitic nematodes cause profound economic losses to global agriculture with the obligate sedentary endoparasitic varieties; amongst them the cyst and Root Knot Nematode (RKN) species are the most damaging. Meloidogyne graminicola is a RKN mainly found in the monocotyledous plants. In the compatible interaction with Oryza sativa, M. graminicola induces the characteristic formation of hook-like galls resulting from the redifferentiation of root cells into multinucleate giant cells. In order to understand the global transcriptome changes occurring during infection, several recent microarray studies on root knots have demonstrated complex changes in host plant gene expression in response to infection. However, to our knowledge, all these transcriptome studies were performed on dicotyledous plants. A histological study enabled us to observe hyperplasia and hypertrophy of the surrounding cells leading to the formation of hook-like galls. We also investigated the plant response to M. graminicola by carrying out a global analysis of gene expression during gall formation in rice, using giant cell-enriched root tissues at an early stage (2dpi) and a latter stage (4dpi) of gall development.-Oryza sativa (var. Nipponbare) seedlings were grown on 6 cm3 SAP substrate completed with diluted Hoaglands solution (Reversat et al., 1999). Culture units were placed in a growth chamber illuminated with fluorescent tubes 9/24 h and maintained at 23°C for 5 days before being inoculated with a 100 J2-stage juveniles M. graminicola. One day after inoculation (dai), the rice seedlings were immersed in de-ionised water to remove all J2s that had not penetrated the roots and allowing synchronization of the infection. Each seedling was transferred to a hydroponic mini chamber (Reversat et al., 2004). Sampling was performed at 2 and 4 dai and each of them contained galls from 70 infected plants, they were then hand-dissected, frozen in liquid-nitrogen and stored at -80°C. As reference samples, uninfected meristematic root fragments were dissected from seedlings grown under the same conditions. Each sample was replicated 3 times. Keywords: normal vs disease comparison,time course

Dataset Information

Identification of GCC-box and TCC-box motifs in the promoters of differentially expressed genes in rice (Oryza sativa L.): Experimental and computational approaches.

Publications

Identification of GCC-box and TCC-box motifs in the promoters of differentially expressed genes in rice (Oryza sativa L.): Experimental and computational approaches.

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