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    Global transcriptional analysis reveals unique and shared responses in Arabidopsis thaliana exposed to combined drought and pathogen stress
    (Frontiers Media S.A., 2016) Gupta, Aarti; Sarkar, Ananda K.; Senthil-Kumar, Muthappa
    With frequent fluctuations in global climate, plants are exposed to co-occurring drought and pathogen infection and this combination adversely affects plant survival. In the past, some studies indicated that morpho-physiological responses of plants to the combined stress are different from the individual stressed plants. However, interaction of drought stressed plants with pathogen has not been widely studied at molecular level. Such studies are important to understand the defense pathways that operate as part of combined stress tolerance mechanism. In this study, Arabidopsis thaliana was exposed to individual drought stress, Pseudomonas syringae pv tomato DC3000 (Pst DC3000) infection and their combination. Using Affymetrix WT gene 1.0 ST array, global transcriptome profiling of leaves under individual drought stress and pathogen infection was compared with their combination. The results obtained from pathway mapping (KAAS and MAPMAN) demonstrated the modulation in defense pathways in A. thaliana under drought and host pathogen Pst DC3000 infection. Further, our study revealed ‘tailored’ responses under combined stress and the time of occurrence of each stress during their concurrence has showed differences in transcriptome profile. Our results from microarray and RT-qPCR revealed unique regulation of 20 novel genes exclusively during the stress interaction. This study indicates that plants exposed to concurrent drought and pathogen stress experience a new state of stress. Thus, under frequently changing climatic conditions each combination of stressor and their timing defines the plant responses and should thus be studied explicitly.
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    Functional relationship of GBF1 with HY5 and HYH in genome-wide gene expression in Arabidopsis
    (Springer, 2016) Ram, Hathi; Jain, Mukesh; Singh, Aparna; Chattopadhyay, Sudip
    Transcriptional networks play important roles in the regulation of biological processes through coordinated activation or repression of downstream target genes. Arabidopsis bZIP transcription factors, GBF1, HY5, and HYH, interact and heterodimerize with each other to form the regulatory network in photomorphogenesis. The genome-wide direct target genes of GBF1 and the roles of HY5 and HYH in controlling GBF1’s genome-wide DNA binding ability have been shown earlier. However, the GBF1 regulated genes at global scale, and how HY5 and HYH modulate GBF1-mediated genome-wide gene expression remain unknown. Here, we report the genome-wide gene expression profile in gbf1, gbf1 hy5, and gbf1 hyh mutants. Our results suggest that HY5 and HYH antagonistically regulate GBF1-mediated global gene expression. We validated the microarray analysis with independent qPCR analyses. Functional analysis of GBF1-regulated genes validates previously known roles of GBF1 in important biological processes. Furthermore, the data also highlight possible novel role of GBF1 in several other biological processes. The previous ChIP-chip results and this transcriptome data together demonstrate the complex transcriptional regulatory mechanism of these transcription factors, GBF1, HY5, and HYH, in photomorphogenesis.
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    Genome-wide transcriptome modulation in rice transgenic lines expressing engineered mitogen activated protein kinase kinase 6
    (Taylor & Francis Group, 2014) Kumar, Kundan; Sinha, Alok Krishna
    Mitogen activated protein kinase kinase (MAPKK) is the central module of MAPK cascade and also point of signal integration and divergence. To investigate the regulatory role of OsMKK6, the regulon of genes controlled by OsMKK6 was constructed by microarray analysis between constitutively activated overexpressing transgenic lines and the wild type rice. Regulated genes were identified in overexpressed constitutively activated OsMKK6 and they were further subdivided on the basis of functional categories, viz. transcription, metabolism, signaling, defense and unknown function. These findings suggest the possible physiological role of OsMKK6 in modulating gene expression and signaling pathways during different stresses.
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    Expanding frontiers in plant transcriptomics in aid of functional genomics and molecular breeding
    (John Wiley & Sons Ltd, 2014) Agarwal, Pinky; Parida, Swarup K.; Mahto, Arunima; Das, Sweta; Mathew, Iny Elizebeth; Malik, Naveen; Tyagi, Akhilesh K.
    The transcript pool of a plant part, under any given condition, is a collection of mRNAs that will pave the way for a biochemical reaction of the plant to stimuli. Over the past decades, transcriptome study has advanced from Northern blotting to RNA sequencing (RNA-seq), through other techniques, of which real-time quantitative polymerase chain reaction (PCR) and microarray are the most significant ones. The questions being addressed by such studies have also matured from a solitary process to expression atlas and marker-assisted genetic enhancement. Not only genes and their networks involved in various developmental processes of plant parts have been elucidated, but also stress tolerant genes have been highlighted. The transcriptome of a plant with altered expression of a target gene has given information about the downstream genes. Marker information has been used for breeding improved varieties. Fortunately, the data generated by transcriptome analysis has been made freely available for ample utilization and comparison. The review discusses this wide variety of transcriptome data being generated in plants, which includes developmental stages, abiotic and biotic stress, effect of altered gene expression, as well as comparative transcriptomics, with a special emphasis on microarray and RNA-seq. Such data can be used to determine the regulatory gene networks, which can subsequently be utilized for generating improved plant varieties.
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    Transcriptome analysis of rin mutant fruit and in silico analysis of promoters of differentially regulated genes provides insight into LeMADS-RIN-regulated ethylene-dependent as well as ethylene-independent aspects of ripening in tomato
    (Springer Science, 2012) Kumar, Rahul; Sharma, Manoj K.; Kapoor, Sanjay; Tyagi, Akhilesh K.; Sharma, Arun K.
    A thorough understanding of molecular mechanisms underlying ripening is the prerequisite for genetic manipulation of fruits for better shelf-life and nutritional quality. Mutation in LeMADS-RIN, a MADS-box gene, leads to non-ripening phenotype of rin fruits in tomato. Characterization of ripening-inhibitor (rin) mutant has elucidated important role of ethylene in the regulation of climacteric fruit ripening. A complete understanding of this mutation will unravel novel genetic regulatory mechanisms involved in fruit ripening. In this study, fruit transcriptomes of two genotypes, including a cultivated Indian cultivar Solanum lycopersicum cv. Pusa Ruby and a homozygous line harboring the rin mutation (LA1795) were compared to get better insight into RIN-regulated ethylene-dependent and ethylene-independent events during ripening. Cluster analysis of ripening-related genes indicated a major shift in their expression profiles in rin mutant fruit. A total of 112 genes, exhibiting expression patterns similar to that of LeMADS-RIN in wild-type fruits, showed down regulation of expression in the rin mutant. In silico analysis of putative promoters of these genes for the presence of CArG box along with ERE and ethylene inducibility of these genes revealed that genes lacking CArG box in their regulatory regions could be indirectly regulated by LeMADS-RIN. New regulators of ethylene-dependent aspect of ripening were also identified. In this study, we have made an attempt to distinguish between ethylene-dependent and ethylene-independent aspects of ripening, which will be useful for developing strategies to improve fruit-related agronomic traits in tomato and other crops.
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    Modulation of transcription factor and metabolic pathway genes in response to water-deficit stress in rice
    (Springer Science, 2011) Ray, Swatismita; Dansana, Prasant K.; Giri, Jitender; Deveshwar, Priyanka; Arora, Rita; Agarwal, Pinky; Khurana, Jitendra P.; Kapoor, Sanjay; Tyagi, Akhilesh K.
    Water-deficit stress is detrimental for rice growth, development, and yield. Transcriptome analysis of 1-week-old rice (Oryza sativa L. var. IR64) seedling under water-deficit stress condition using Affymetrix 57 K GeneChip® has revealed 1,563 and 1,746 genes to be up- and downregulated, respectively. In an effort to amalgamate data across laboratories, we identified 5,611 differentially expressing genes under varying extrinsic water-deficit stress conditions in six vegetative and one reproductive stage of development in rice. Transcription factors (TFs) involved in ABA-dependent and ABA-independent pathways have been found to be upregulated during water-deficit stress. Members of zinc-finger TFs namely, C₂H₂, C₂C₂, C₃H, LIM, PHD, WRKY, ZF-HD, and ZIM, along with TF families like GeBP, jumonji, MBF1 and ULT express differentially under water-deficit conditions. NAC (NAM, ATAF and CUC) TF family emerges to be a potential key regulator of multiple abiotic stresses. Among the 12 TF genes that are co-upregulated under water-deficit, salt and cold stress conditions, five belong to the NAC TF family. We identified water-deficit stress-responsive genes encoding key enzymes involved in biosynthesis of osmoprotectants like polyols and sugars; amino acid and quaternary ammonium compounds; cell wall loosening and structural components; cholesterol and very long chain fatty acid; cytokinin and secondary metabolites. Comparison of genes responsive to water-deficit stress conditions with genes preferentially expressed during panicle and seed development revealed a significant overlap of transcriptome alteration and pathways.
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    Genome-wide identification of novel internal control genes for normalization of gene-expression during various stages of development in rice
    (Elsevier B.V., 2009) Jain, Mukesh
    An internal control gene with highly uniform expression throughout the experimental conditions is required for accurate and reliable gene expression results. However, no gene can serve as a universal internal control, implying the need for identification of most suitable internal control gene(s) for experimental conditions being analyzed. In an earlier study, we identified UBQ5 and eEF1-α as the most suitable internal control genes by validating the expression of ten housekeeping genes in rice. The aim of this study was to identify the most suitable novel internal control genes at the whole genome level for normalization of gene expression during various developmental stages in rice, the model crop plant. A systematic analysis of the whole genome microarray data for various stages of vegetative and reproductive development in rice was performed. Several novel internal control genes have been identified, which display highly uniform expression levels in all the developmental stages analyzed and outperform many traditional internal control genes. The expression of these genes ranges from low to very high, which make them suitable for normalization of gene expression over a wide range of transcript levels. The expression stability of the novel genes identified has also been validated by geNORM and NormFinder softwares. The novel genes with highly uniform expression identified in this study can be used for more accurate normalization of transcript levels during various developmental stages in rice and should enable the detection of biologically significant changes in gene expression.