Regulation of Nuclear Localization of Signaling Proteins by Cytokinin

Cytokinins are a class of mitogenic plant hormones that play an important role in most aspects of plant development, including shoot and root growth, vascular and photomorphogenic development and leaf senescence. A model for cytokinin perception and signaling has emerged that is similar to bacterial two-component phosphorelays. In this model, binding of cytokinin to the extracellular domain of the Arabidopsis histidine kinase (AHKs) receptors induces autophosphorylation within the intracellular histidine-kinase domain. The phosphoryl group is subsequently transferred to cytosolic Arabidopsis histidine phosphotransfer proteins (AHPs), which have been suggested to translocate to the nucleus in response to cytokinin treatment, where they then transfer the phosphoryl group to nuclear-localized response regulators (Type-A and Type-B ARRs). We examined the effects of cytokinin on AHP subcellular localization in Arabidopsis and, contrary to expectations, the AHPs maintained a constant nuclear/cytosolic distribution following cytokinin treatment. Furthermore, mutation of the conserved phosphoacceptor histidine residue of the AHP, as well as disruption of multiple cytokinin signaling elements, did not affect the subcellular localization of the AHP proteins. Finally, we present data indicating that AHPs maintain a nuclear/cytosolic distribution by balancing active transport into and out of the nucleus. Our findings suggest that the current models indicating relocalization of AHP protein into the nucleus in response to cytokinin are incorrect. Rather, AHPs actively maintain a consistent nuclear/cytosolic distribution regardless of the status of the cytokinin response pathway

Windstein – Château du Nouveau-Windstein

Après le dégagement et la mise en valeur de la barbacane à l’entrée du Nouveau-Windstein, il est proposé de poursuivre le dégagement des abords immédiats du château dans l’objectif de : restituer la logique défensive du château en dégageant la base du bastion côté nord. Ce bastion constitue le vis-à-vis de la barbacane pour la défense de l’esplanade et constitue également la défense du chemin d’accès et de la porte cochère à l’entrée de l’esplanade (cf. travaux de 2014). restituer le cheminem..

Windstein – Château du Nouveau-Windstein

Après les sondages archéologiques aux abords immédiats de la barbacane, côté esplanade, réalisés entre 2009 et 2014, les bénévoles des veilleurs du Nouveau-Windstein ont réalisé le sondage au pied de la barbacane côté vallée, à l’extérieur de l’enceinte du château. L’encadrement scientifique est assuré par le service régional de l’archéologie. Les principaux objectifs archéologiques étaient la restitution de la logique défensive du château dans sa période tardive d’occupation et l’analyse de ..

High dimethylsulfide photolysis rates in nitrate-rich Antarctic waters

Author Posting. © American Geophysical Union, 2004. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Geophysical Research Letters 31 (2004): L11307, doi:10.1029/2004GL019863.The photochemistry of dimethylsulfide (DMS) was examined in the Southern Ocean to assess its impact on the biogeochemical dynamics of DMS in Antarctic waters. Very high DMS photolysis rate constants (0.16–0.23 h−1) were observed in surface waters exposed to full sunlight. DMS photolysis rates increased linearly with added nitrate concentrations, and 35% of the DMS loss in unamended samples was attributed to the photochemistry of ambient nitrate (29 μM). Experiments with optical filters showed that the UV-A band of sunlight (320–400 nm) accounted for ~65% of DMS photolysis suggesting that dissolved organic matter was the main photosensitizer for DMS photolysis. During the austral spring, DMS photolysis was the dominant loss mechanism under non-bloom and non-ice cover conditions owing to the high doses and deep penetration of UV radiation in the water column, low observed microbial consumption rates, and high in situ nitrate concentrations.This work was supported by NSF (OPP- 0230499, DJK; OPP-0230497, RPK)

The Promise of Anti-Idiotype Revisited

The promise of idiotype-based therapeutics has been disappointing forcing a new look at the concept and its potential to generate an effective approach for immunotherapy. Here, the idiotype network theory is revisited with regard to the development of efficacious anti-idiotype vaccines. The experience of polyclonal anti-Idiotype reagents in animal models as well as an understanding of the immune response in humans lends to the proposition that polyclonal anti-Idiotype vaccines will be more effective compared to monoclonal-based anti-Idiotype vaccines. This novel strategy can be adapted in Biotech-standard production of therapeutic antibodies

indCAPS: A tool for designing screening primers for CRISPR/Cas9 mutagenesis events

Genetic manipulation of organisms using CRISPR/Cas9 technology generally produces small insertions/deletions (indels) that can be difficult to detect. Here, we describe a technique to easily and rapidly identify such indels. Sequence-identified mutations that alter a restriction enzyme recognition site can be readily distinguished from wild-type alleles using a cleaved amplified polymorphic sequence (CAPS) technique. If a restriction site is created or altered by the mutation such that only one allele contains the restriction site, a polymerase chain reaction (PCR) followed by a restriction digest can be used to distinguish the two alleles. However, in the case of most CRISPR-induced alleles, no such restriction sites are present in the target sequences. In this case, a derived CAPS (dCAPS) approach can be used in which mismatches are purposefully introduced in the oligonucleotide primers to create a restriction site in one, but not both, of the amplified templates. Web-based tools exist to aid dCAPS primer design, but when supplied sequences that include indels, the current tools often fail to suggest appropriate primers. Here, we report the development of a Python-based, species-agnostic web tool, called indCAPS, suitable for the design of PCR primers used in dCAPS assays that is compatible with indels. This tool should have wide utility for screening editing events following CRISPR/Cas9 mutagenesis as well as for identifying specific editing events in a pool of CRISPR-mediated mutagenesis events. This tool was field-tested in a CRISPR mutagenesis experiment targeting a cytokinin receptor (AHK3) in Arabidopsis thaliana. The tool suggested primers that successfully distinguished between wild-type and edited alleles of a target locus and facilitated the isolation of two novel ahk3 null alleles. Users can access indCAPS and design PCR primers to employ dCAPS to identify CRISPR/Cas9 alleles at http://indcaps.kieber.cloudapps.unc.edu/

Determination of Photochemically Produced Carbon Dioxide in Seawater

An analytical system was developed to determine photochemically produced carbon dioxide in marine waters. Our system was designed to measure low levels of carbon dioxide by maintaining a closed system to prevent atmospheric contamination during sample preparation, irradiation, and analysis. To detect low levels of photoproduced carbon dioxide in seawater, background dissolved inorganic carbon (DIC) was removed before irradiation. To strip out DIC, samples were acidified to pH 3.0 (converting DIC to carbon dioxide) and bubbled with low carbon dioxide air. The pH was then readjusted back to the original value, and the resulting low-DIC seawater samples were transferred pneumatically to air-tight quartz tubes for irradiation. During analysis, samples were pneumatically transferred to a sample loop, injected, and acidified. Carbon dioxide was then stripped out, dried, and carried to a nondispersive infrared carbon dioxide analyzer. Calibration was done with a series of low concentration aqueous carbonate standards (0.05 to 3 μmol L-1). The detection limit, defined as the concentration corresponding to three times the standard deviation of the experimental blank (i.e., DIC-stripped seawater), was similar to ~60 nmol L-1. Method precision was largely dependent on the agreement between multiple injections from the same tube (\u3c ± 2% relative standard deviation [RSD]) and the reproducibility between different tubes (±3% RSD). This method was used to measure carbon dioxide photoproduction in a variety of waters (e.g., estuarine, lake) including the first direct measurements in marine waters