Journal of Proteomics & Bioinformatics- Open Access www.omicsonline.com Research Article JPB/Vol.2/August 2009 In Silico Identification of Putative Proton Binding Sites of a Plasma Membrane H +-ATPase Isoform of Arabidopsis Thaliana, AHA1

jpb.1000095 Copyright: © 2009 Kumar S, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The plasma membrane potential and secondary transport systems in all eukaryotes are energized by the activity of P-type ATPase membrane proteins: H + ATPase (the proton pump) in plants and fungi and Na +, K +- ATPase (the sodium-potassium pump) in animals. The overall shape of proton pumps has been revealed by electron microscopy. The crystal structure of AHA2, a plasma membrane H +-ATPase isoform of Arabidopsis thaliana, by X-ray crystallography at 3.6 Å was available. The isoform is expressed mainly in root. In the present study homology modeling along with transmembrane topology predictions has been used to build the atomic model of AHA1, another plasma membrane H +-ATPase isoform of Arabidopsis thaliana expressing in both root and shoot. AHA2 was used as the template. The homology modeling was done using the MODELLER9v2 software. The model energy was minimized by applying molecular mechanics method. The root mean square deviation (RMSD) for C

Biochemical and molecular characterisation of exogenous cytokinin application on grain filling in rice

Abstract Background Poor filling of grains in the basal spikelets of large size panicles bearing numerous spikelets has been a major limitation in attempts to increase the rice production to feed the world’s increasing population. Considering that biotechnological intervention could play important role in overcoming this limitation, the role of cytokinin in grain filling was investigated based on the information on cell proliferating potential of the hormone and reports of its high accumulation in immature seeds. Results A comparative study considering two rice varieties differing in panicle compactness, lax-panicle Upahar and compact-panicle OR-1918, revealed significant difference in grain filling, cytokinin oxidase (CKX) activity and expression, and expression of cell cycle regulators and cytokinin signaling components between the basal and apical spikelets of OR-1918, but not of Upahar. Exogenous application of cytokinin (6-Benzylaminopurine, BAP) to OR-1918 improved grain filling significantly, and this was accompanied by a significant decrease in expression and activity of CKX, particularly in the basal spikelets where the activity of CKX was significantly higher than that in the apical spikelets. Cytokinin application also resulted in significant increase in expression of cell cycle regulators like cyclin dependent kinases and cyclins in the basal spikelets that might be facilitating cell division in the endosperm cells by promoting G1/S phase and G2/M phase transition leading to improvement in grain filling. Expression studies of type-A response regulator (RR) component of cytokinin signaling indicated possible role of OsRR3, OsRR4 and OsRR6 as repressors of CKX expression, much needed for an increased accumulation of CK in cells. Furthermore, the observed effect of BAP might not be solely because of it, but also because of induced synthesis of trans-zeatin (tZ) and N6-(Δ2-isopentenyl)adenine (iP), as reflected from accumulation of tZR (tZ riboside) and iPR (iP riboside), and significantly enhanced expression of an isopentenyl transferase (IPT) isoform. Conclusion The results suggested that seed-specific overexpression of OsRR4 and OsRR6, and more importantly of IPT9 could be an effective biotechnological intervention towards improving the CK level of the developing caryopses leading to enhanced grain filling in rice cultivars bearing large panicles with numerous spikelets, and thereby increasing their yield potential

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Transcriptome Analysis of the Response to NaCl in <i>Suaeda maritima</i> Provides an Insight into Salt Tolerance Mechanisms in Halophytes

<div><p>Although salt tolerance is a feature representative of halophytes, most studies on this topic in plants have been conducted on glycophytes. Transcriptome profiles are also available for only a limited number of halophytes. Hence, the present study was conducted to understand the molecular basis of salt tolerance through the transcriptome profiling of the halophyte <i>Suaeda maritima</i>, which is an emerging plant model for research on salt tolerance. Illumina sequencing revealed 72,588 clustered transcripts, including 27,434 that were annotated using BLASTX. Salt application resulted in the 2-fold or greater upregulation of 647 genes and downregulation of 735 genes. Of these, 391 proteins were homologous to proteins in the COGs (cluster of orthologous groups) database, and the majorities were grouped into the poorly characterized category. Approximately 50% of the genes assigned to MapMan pathways showed homology to <i>S</i>. <i>maritima</i>. The majority of such genes represented transcription factors. Several genes also contributed to cell wall and carbohydrate metabolism, ion relation, redox responses and G protein, phosphoinositide and hormone signaling. Real-time PCR was used to validate the results of the deep sequencing for the most of the genes. This study demonstrates the expression of protein kinase C, the target of diacylglycerol in phosphoinositide signaling, for the first time in plants. This study further reveals that the biochemical and molecular responses occurring at several levels are associated with salt tolerance in <i>S</i>. <i>maritima</i>. At the structural level, adaptations to high salinity levels include the remodeling of cell walls and the modification of membrane lipids. At the cellular level, the accumulation of glycinebetaine and the sequestration and exclusion of Na⁺ appear to be important. Moreover, this study also shows that the processes related to salt tolerance might be highly complex, as reflected by the salt-induced enhancement of transcription factor expression, including hormone-responsive factors, and that this process might be initially triggered by G protein and phosphoinositide signaling.</p></div

Identification and expression analysis of miRNAs and elucidation of their role in salt tolerance in rice varieties susceptible and tolerant to salinity.

Soil salinization is a serious problem for cultivation of rice, as among cereals rice is the most salt sensitive crop, and more than 40% of the total agricultural land amounting to approximately 80 million ha the world over is salt affected. Salinity affects a plant in a varieties of ways, including ion toxicity, osmotic stress and oxidative damage. Since miRNAs occupy the top place in biochemical events determining a trait, understanding their role in salt tolerance is highly desirable, which may allow introduction of the trait in the rice cultivars of choice through biotechnological interventions. High throughput sequencing of sRNAs in the root and shoot tissues of the seedlings of the control and NaCl treated Pokkali, a salt-tolerant rice variety, identified 75 conserved miRNAs and mapped 200 sRNAs to the rice genome as novel miRNAs. Expression of nine novel miRNAs and two conserved miRNAs were confirmed by Northern blotting. Several of both conserved and novel miRNAs that expressed differentially in root and/or shoot tissues targeted transcription factors like AP2/EREBP domain protein, ARF, NAC, MYB, NF-YA, HD-Zip III, TCP and SBP reported to be involved in salt tolerance or in abiotic stress tolerance in general. Most of the novel miRNAs expressed in the salt tolerant wild rice Oryza coarctata, suggesting conservation of miRNAs in taxonomically related species. One of the novel miRNAs, osa-miR12477, also targeted L-ascorbate oxidase (LAO), indicating build-up of oxidative stress in the plant upon salt treatment, which was confirmed by DAB staining. Thus, salt tolerance might involve miRNA-mediated regulation of 1) cellular abundance of the hormone signaling components like EREBP and ARF, 2) synthesis of abiotic stress related transcription factors, and 3) antioxidative component like LAO for mitigation of oxidative damage. The study clearly indicated importance of osa-miR12477 regulated expression of LAO in salt tolerance in the plant

Primary assembly statistics of the control and 2.0% NaCl-treated (9 h) <i>S</i>. <i>maritima</i> libraries generated using Trinity assembler.

<p>Primary assembly statistics of the control and 2.0% NaCl-treated (9 h) <i>S</i>. <i>maritima</i> libraries generated using Trinity assembler.</p

Secondary assembly statistics of the control and 2.0% NaCl-treated (9 h) <i>S</i>. <i>maritima</i> libraries following CD-HIT EST analysis.

<p>Secondary assembly statistics of the control and 2.0% NaCl-treated (9 h) <i>S</i>. <i>maritima</i> libraries following CD-HIT EST analysis.</p

Changes in the expression of transcription factors in <i>S</i>. <i>maritima</i> in response to NaCl application.

<p>The grey bars represent the transcription factors that showed significant changes (2-fold or greater and/or changes significant at <i>p</i> ≤ 0.05), either upregulation or downregulation, based on the sequencing results in response to NaCl application. The corresponding black bars show expression levels as determined by qPCR, expressed as fold-changes relative to expression in the controls, in response to the NaCl application. The individual black bars, representing the qPCR data, are the means ± SD of six measurements (three technical replicates each for two biological samples). *, ** and *** next to a column indicate significant changes in expression at <i>p</i> ≤ 0.05, <i>p</i> ≤ 0.01 and <i>p</i> ≤ 0.001, respectively. Ns = not significant.</p

Pie chart showing the influence of NaCl on the distribution of reads among the three GO components, i.e., biological process, molecular function and cellular component.

<p>Pie chart showing the influence of NaCl on the distribution of reads among the three GO components, i.e., biological process, molecular function and cellular component.</p