The sucrose non-fermenting 1-related kinase 2 gene SAPK9 improves drought tolerance and grain yield in rice by modulating cellular osmotic potential, stomatal closure and stress-responsive gene expression

Abstract Background Family members of sucrose non-fermenting 1-related kinase 2 (SnRK2), being plant-specific serine/threonine protein kinases, constitute the central core of abscisic acid (ABA)-dependent and ABA-independent signaling pathways, and are key regulators of abiotic stress adaptation in plants. We report here the functional characterization of SAPK9 gene, one of the 10 SnRK2s of rice, through developing gain-of-function and loss-of-function phenotypes by transgenesis. Results The gene expression profiling revealed that the abundance of single gene-derived SAPK9 transcript was significantly higher in drought-tolerant rice genotypes than the drought-sensitive ones, and its expression was comparatively greater in reproductive stage than the vegetative stage. The highest expression of SAPK9 gene in drought-tolerant Oryza rufipogon prompted us to clone and characterise the CDS of this allele in details. The SAPK9 transcript expression was found to be highest in leaf and upregulated during drought stress and ABA treatment. In silico homology modelling of SAPK9 with Arabidopsis OST1 protein showed the bilobal kinase fold structure of SAPK9, which upon bacterial expression was able to phosphorylate itself, histone III and OsbZIP23 as substrates in vitro. Transgenic overexpression (OE) of SAPK9 CDS from O. rufipogon in a drought-sensitive indica rice genotype exhibited significantly improved drought tolerance in comparison to transgenic silencing (RNAi) lines and non-transgenic (NT) plants. In contrast to RNAi and NT plants, the enhanced drought tolerance of OE lines was concurrently supported by the upgraded physiological indices with respect to water retention capacity, soluble sugar and proline content, stomatal closure, membrane stability, and cellular detoxification. Upregulated transcript expressions of six ABA-dependent stress-responsive genes and increased sensitivity to exogenous ABA of OE lines indicate that the SAPK9 is a positive regulator of ABA-mediated stress signaling pathways in rice. The yield-related traits of OE lines were augmented significantly, which resulted from the highest percentage of fertile pollens in OE lines when compared with RNAi and NT plants. Conclusion The present study establishes the functional role of SAPK9 as transactivating kinase and potential transcriptional activator in drought stress adaptation of rice plant. The SAPK9 gene has potential usefulness in transgenic breeding for improving drought tolerance and grain yield in crop plants.http://deepblue.lib.umich.edu/bitstream/2027.42/134605/1/12870_2016_Article_845.pd

PRC2 represses transcribed genes on the imprinted inactive X chromosome in mice

Abstract Background Polycomb repressive complex 2 (PRC2) catalyzes histone H3K27me3, which marks many transcriptionally silent genes throughout the mammalian genome. Although H3K27me3 is associated with silenced gene expression broadly, it remains unclear why some but not other PRC2 target genes require PRC2 and H3K27me3 for silencing. Results Here we define the transcriptional and chromatin features that predict which PRC2 target genes require PRC2/H3K27me3 for silencing by interrogating imprinted mouse X-chromosome inactivation. H3K27me3 is enriched at promoters of silenced genes across the inactive X chromosome. To abrogate PRC2 function, we delete the core PRC2 protein EED in F1 hybrid trophoblast stem cells (TSCs), which undergo imprinted inactivation of the paternally inherited X chromosome. Eed –/– TSCs lack H3K27me3 and Xist lncRNA enrichment on the inactive X chromosome. Despite the absence of H3K27me3 and Xist RNA, only a subset of the inactivated X-linked genes is derepressed in Eed –/– TSCs. Unexpectedly, in wild-type (WT) TSCs these genes are transcribed and are enriched for active chromatin hallmarks on the inactive-X, including RNA PolII, H3K27ac, and H3K36me3, but not the bivalent mark H3K4me2. By contrast, PRC2 targets that remain repressed in Eed –/– TSCs are depleted for active chromatin characteristics in WT TSCs. Conclusions A comparative analysis of transcriptional and chromatin features of inactive X-linked genes in WT and Eed –/– TSCs suggests that PRC2 acts as a brake to prevent induction of transcribed genes on the inactive X chromosome, a mode of PRC2 function that may apply broadly.https://deepblue.lib.umich.edu/bitstream/2027.42/136651/1/13059_2017_Article_1211.pd

Prediction-based protein engineering of domain I of Cry2A entomocidal toxin of Bacillus thuringiensis for the enhancement of toxicity against lepidopteran insects

Issues relating to sustenance of the usefulness of genetically modified first generation Bt crop plants in the farmer’s field are of great concern for crop scientists. Additional biotechnological strategies need to be in place to safeguard the possibility for yield loss of Bt crop by other lepidopteran insects that are insensitive to the Cry1A toxin, and also against the possibility for emergence of resistant insects. In this respect, Cry2A toxin has figured as a prospective candidate to be the second toxin to offer the required protection along with Cry1A. In the present study, the entomocidal potency of Cry2A toxin was enhanced through knowledge-based protein engineering of the toxin molecule. Deletion of 42 amino acid residues from the N-terminal end of the peptide followed by the replacement of Lys residues by nonpolar amino acids in the putative transmembrane region including the introduction of Pro resulted in a 4.1–6.6-fold increase in the toxicity of the peptide against three major lepidopteran insect pests of crop plants

Differentiation-dependent Requirement of Tsix long non-coding RNA in Imprinted X-chromosome Inactivation

Imprinted X-inactivation is a paradigm of mammalian transgenerational epigenetic regulation resulting in silencing of genes on the paternally-inherited X-chromosome. The pre-programmed fate of the X-chromosomes is thought to be controlled in cis by the parent-of-origin-specific expression of two long non-coding RNAs, Tsix and Xist, in mice. Exclusive expression of Tsix from the maternal–X has implicated it as the instrument through which the maternal germline prevents inactivation of the maternal–X in the offspring. Here, we show that Tsix is dispensable for inhibiting Xist and X-inactivation in the early embryo and in cultured stem cells of extra-embryonic lineages. Tsix is instead required to prevent Xist expression as trophectodermal progenitor cells differentiate. Despite induction of wild-type Xist RNA and accumulation of histone H3-K27me3, many Tsix-mutant X-chromosomes fail to undergo ectopic X-inactivation. We propose a novel model of lncRNA function in imprinted X-inactivation that may also apply to other genomically imprinted loci

Data Throughput of Wireless Network for Fire Alarms

Import 22/07/2015Tato bakalářská práce se zabývá ostravskou hasičskou sítí, propustností, rušením a návrhem na vylepšení sítě z hlediska datové propustnosti. Analýza datové propustnosti byla provedena pomoci vlastního programu napsaného v C#. Pomocí USB tuneru Rafael Micro R820T s čipsetem RTL2832U a počítačem s operačním systémem Ubuntu 14.04, na kterém byly nainstalován software Librtlsdr, GNU radio GQRX , Teamviewer a Kazam. Těmito programy byly sledovány vstupní kmitočty převaděčů, které neodhalily žádné rušení. Dále byly vypsány možné vlivy teoretického rušení. Následně byly vymyšleny dvě teoreticky zlepšené varianty systému. První se zabývá obousměrným přenosem, kdy koncové vysílače přijímají zprávu o potvrzení přijetí z převaděče a druhá přidáním dalšího převaděče, který by se při správném umístění, které by bylo na výškové budově domova sester. Hlavní výhodou tohoto řešení je větší pokrytí oblasti. Oba tyto návrhy mají lepší vlastnosti v oblasti datové propustnosti.The bachelor thesis deals with the fire-fighting net in Ostrava, it's permeability, disturbance and improvement proposal for this net from the point of view of data permeability. Analysis of data permeability was made by own programme wrote in C#. Disturbance was watched by USB tuner Rafael Micro R820T with chipset RTL2832U and with computer with operating system Ubuntu 14.04. On Ubuntu was install a software Librtlsdr, GNU radio GQRX, Teamviewer and Kazam. But the disturbance was not found. The list of the theroretical influences on disturbance was made. Two theoretical better options were invented. The first one deals with two-way transfer and the second one proposes additional convertor. These suggestions have better properties in the field of data permeability.440 - Katedra telekomunikační technikydobř

A Primary Role for the Tsix lncRNA in Maintaining Random X-Chromosome Inactivation

Differentiating pluripotent epiblast cells in eutherians undergo random X-inactivation, which equalizes X-linked gene expression between the sexes by silencing one of the two X-chromosomes in females. Tsix RNA is believed to orchestrate the initiation of X-inactivation, influencing the choice of which X remains active by preventing expression of the antisense Xist RNA, which is required to silence the inactive-X. Here we profile X-chromosome activity in Tsix-mutant (XΔTsix) mouse embryonic epiblasts, epiblast stem cells, and embryonic stem cells. Unexpectedly, we find that Xist is stably repressed on the XΔTsix in both sexes in undifferentiated epiblast cells in vivo and in vitro, resulting in stochastic X-inactivation in females despite Tsix-heterozygosity. Tsix is instead required to silence Xist on the active-X as epiblast cells differentiate in both males and females. Thus, Tsix is not required at the onset of random X-inactivation; instead, it protects the active-X from ectopic silencing once X-inactivation has commenced

Additional file 5: Figure S3. of The sucrose non-fermenting 1-related kinase 2 gene SAPK9 improves drought tolerance and grain yield in rice by modulating cellular osmotic potential, stomatal closure and stress-responsive gene expression

Amino acid sequence alignment of the isolated SAPK9 CDS from wild rice Oryza rufipogon (accession no. KT387673) with the reported sequence of japonica rice cultivar (accession no.AB125310). (TIF 487 kb

Additional file 2: Figure S5. of The sucrose non-fermenting 1-related kinase 2 gene SAPK9 improves drought tolerance and grain yield in rice by modulating cellular osmotic potential, stomatal closure and stress-responsive gene expression

Schematic representation of the genetic constructs based on pCAMBIA1301 plasmid used for Agrobacterium-mediated transformation of drought-sensitive indica rice cultivar IR20. (A) The gene overexpression (OE) construct of SAPK9 carrying 1086 bp CDS from O. rufipogon. The developed transgenic rice lines were designated as SAOE#1, 2, 3 etc. (B) The RNAi-mediated gene silencing (RNAi) construct of endogenous SAPK9 gene. The 605 bp 5′-part of SAPK9 CDS from O. rufipogon was cloned in sense and antisense orientation flanking an arbitrary 200 bp DNA linker. The developed transgenic rice lines were designated as RNAi#1, 2, 3 etc. (TIF 453 kb

Correction: Enhanced Gene Expression Rather than Natural Polymorphism in Coding Sequence of the OsbZIP23 Determines Drought Tolerance and Yield Improvement in Rice Genotypes.

[This corrects the article DOI: 10.1371/journal.pone.0150763.]

Enhanced Gene Expression Rather than Natural Polymorphism in Coding Sequence of the OsbZIP23 Determines Drought Tolerance and Yield Improvement in Rice Genotypes.

Drought is one of the major limiting factors for productivity of crops including rice (Oryza sativa L.). Understanding the role of allelic variations of key regulatory genes involved in stress-tolerance is essential for developing an effective strategy to combat drought. The bZIP transcription factors play a crucial role in abiotic-stress adaptation in plants via abscisic acid (ABA) signaling pathway. The present study aimed to search for allelic polymorphism in the OsbZIP23 gene across selected drought-tolerant and drought-sensitive rice genotypes, and to characterize the new allele through overexpression (OE) and gene-silencing (RNAi). Analyses of the coding DNA sequence (CDS) of the cloned OsbZIP23 gene revealed single nucleotide polymorphism at four places and a 15-nucleotide deletion at one place. The single-copy OsbZIP23 gene is expressed at relatively higher level in leaf tissues of drought-tolerant genotypes, and its abundance is more in reproductive stage. Cloning and sequence analyses of the OsbZIP23-promoter from drought-tolerant O. rufipogon and drought-sensitive IR20 cultivar showed variation in the number of stress-responsive cis-elements and a 35-nucleotide deletion at 5'-UTR in IR20. Analysis of the GFP reporter gene function revealed that the promoter activity of O. rufipogon is comparatively higher than that of IR20. The overexpression of any of the two polymorphic forms (1083 bp and 1068 bp CDS) of OsbZIP23 improved drought tolerance and yield-related traits significantly by retaining higher content of cellular water, soluble sugar and proline; and exhibited decrease in membrane lipid peroxidation in comparison to RNAi lines and non-transgenic plants. The OE lines showed higher expression of target genes-OsRab16B, OsRab21 and OsLEA3-1 and increased ABA sensitivity; indicating that OsbZIP23 is a positive transcriptional-regulator of the ABA-signaling pathway. Taken together, the present study concludes that the enhanced gene expression rather than natural polymorphism in coding sequence of OsbZIP23 is accountable for improved drought tolerance and yield performance in rice genotypes