Function and evolution of channels and transporters in photosynthetic membranes

Chloroplasts from land plants and algae originated from an endosymbiotic event, most likely involving an ancestral photoautotrophic prokaryote related to cyanobacteria. Both chloroplasts and cyanobacteria have thylakoid membranes, harboring pigment-protein complexes that perform the light-dependent reactions of oxygenic photosynthesis. The composition, function and regulation of these complexes have thus far been the major topics in thylakoid membrane research. For many decades, we have also accumulated biochemical and electrophysiological evidence for the existence of solute transthylakoid transport activities that affect photosynthesis. However, research dedicated to molecular identification of the responsible proteins has only recently emerged with the explosion of genomic information. Here we review the current knowledge about channels and transporters from the thylakoid membrane of Arabidopsis thaliana and of the cyanobacterium Synechocystis sp. PCC 6803. No homologues of these proteins have been characterized in algae, although similar sequences could be recognized in many of the available sequenced genomes. Based on phylogenetic analyses, we hypothesize a host origin for most of the so far identified Arabidopsis thylakoid channels and transporters. Additionally, the shift from a non-thylakoid to a thylakoid location appears to have occurred at different times for different transport proteins. We propose that closer control of and provision for the thylakoid by products of the host genome has been an ongoing process, rather than a one-step event. Some of the proteins recruited to serve in the thylakoid may have been the result of the increased specialization of its pigment-protein composition and organization in green plants. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00018-013-1412-3) contains supplementary material, which is available to authorized users

Phylogenetic Analysis of the Thylakoid ATP/ADP Carrier Reveals New Insights into Its Function Restricted to Green Plants

ATP is the common energy currency of cellular metabolism in all living organisms. Most of them synthesize ATP in the cytosol or on the mitochondrial inner membrane, whereas land plants, algae, and cyanobacteria also produce it on the thylakoid membrane during the light-dependent reactions of photosynthesis. From the site of synthesis, ATP is transported to the site of utilization via intracellular membrane transporters. One major type of ATP transporters is represented by the mitochondrial ADP/ATP carrier family. Here we review a recently characterized member, namely the thylakoid ATP/ADP carrier from Arabidopsis thaliana (AtTAAC). Thus far, no orthologs of this carrier have been characterized in other organisms, although similar sequences can be recognized in many sequenced genomes. Protein Sequence database searches and phylogenetic analyses indicate the absence of TAAC in cyanobacteria and its appearance early in the evolution of photosynthetic eukaryotes. The TAAC clade is composed of carriers found in land plants and some green algae, but no proteins from other photosynthetic taxa, such as red algae, brown algae, and diatoms. This implies that TAAC-like sequences arose only once before the divergence of green algae and land plants. Based on these findings, it is proposed that TAAC may have evolved in response to the need of a new activity in higher photosynthetic eukaryotes. This activity may provide the energy to drive reactions during biogenesis and turnover of photosynthetic complexes, which are heterogeneously distributed in a thylakoid membrane system composed of appressed and non-appressed regions

Facilitated Peptide Transport via the Mucosal Epithelium

A hallmark of autoimmunity is the breakdown of tolerance and generation of effector responses against self-antigens. Re-establishment of tolerance in autoimmune disorders was always the most desired treatment option; however, despite many efforts, clinical trials have been largely unsuccessful. This also applies to the generation of oral tolerance, which seems to be a default response type of the mucosa-associated lymphoid tissues to harmless antigens. In this study, we report improved efficacy of oral tolerance induction by coupling antigen with the newly identified mucosal carrier peptide 13C. Antigen coupled to 13C is efficiently taken up in the gastrointestinal tract and could be visualized in cells of the lamina propria. Oral, rectal, or nasal treatment effectively induced the proliferation of antigen-specific T cells with some increase in the frequency of regulatory T cells. In a model of delayed-type hypersensitivity, especially intrarectal tolerization treatment resulted in reduced footpad swelling, demonstrating a moderate tolerogenic effect of mucosal treatment with 13C coupled antigen. Coupling of antigens to a transmucosal carrier, therefore, is a promising tool to improve the efficacy of vaccination via mucosal surfaces

Nanoaperture fabrication via colloidal lithography for single molecule fluorescence analysis

In single molecule fluorescence studies, background emission from labeled substrates often restricts their concentrations to non-physiological nanomolar values. One approach to address this challenge is the use of zero-mode waveguides (ZMWs), nanoscale holes in a thin metal film that physically and optically confine the observation volume allowing much higher concentrations of fluorescent substrates. Standard fabrication of ZMWs utilizes slow and costly E-beam nano-lithography. Herein, ZMWs are made using a self-assembled mask of polystyrene microspheres, enabling fabrication of thousands of ZMWs in parallel without sophisticated equipment. Polystyrene 1 mu m dia. microbeads self-assemble on a glass slide into a hexagonal array, forming a mask for the deposition of metallic posts in the inter-bead interstices. The width of those interstices (and subsequent posts) is adjusted within 100-300 nm by partially fusing the beads at the polystyrene glass transition temperature. The beads are dissolved in toluene, aluminum or gold cladding is deposited around the posts, and those are dissolved, leaving behind an array ZMWs. Parameter optimization and the performance of the ZMWs are presented. By using colloidal self-assembly, typical laboratories can make use of sub-wavelength ZMW technology avoiding the availability and expense of sophisticated clean-room environments and equipment

Overview of the Nordic Seas CARINA data and salinity measurements

Water column data of carbon and carbon relevant hydrographic and hydrochemical parameters from 188 previously non-publicly available cruises in the Arctic, Atlantic, and Southern Ocean have been retrieved and merged into a new database: CARINA (CARbon IN the Atlantic). The data have been subject to rigorous quality control (QC) in order to ensure highest possible quality and consistency. The data for most of the parameters included were examined in order to quantify systematic biases in the reported values, i.e. secondary quality control. Significant biases have been corrected for in the data products, i.e. the three merged files with measured, calculated and interpolated values for each of the three CARINA regions; the Arctic Mediterranean Seas (AMS), the Atlantic (ATL) and the Southern Ocean (SO). With the adjustments the CARINA database is consistent both internally as well as with GLODAP (Key et al., 2004) and is suitable for accurate assessments of, for example, oceanic carbon inventories and uptake rates and for model validation. The Arctic Mediterranean Seas include the Arctic Ocean and the Nordic Seas, and the quality control was carried out separately in these two areas. This contribution provides an overview of the CARINA data from the Nordic Seas and summarises the findings of the QC of the salinity data. One cruise had salinity data that were of questionable quality, and these have been removed from the data product. An evaluation of the consistency of the quality controlled salinity data suggests that they are consistent to at least ±0.005

The Threshold Pion-Photoproduction of Nucleons In The Chiral Quark Model

In this paper, we show that the low energy theorem (LET) of the threshold pion-photoproduction can be fully recovered in the quark model. An essential result of this investigation is that the quark-pion operators are obtained from the effective chiral Lagrangian, and the low energy theorem does not require the constraints on the internal structures of the nucleon. The pseudoscalar quark-pion coupling generates an additional term at order $\mu=m_{\pi}/M$ only in the isospin amplitude $A^{(-)}$. The role of the transitions between the nucleon and the resonance $P_{33}(1232)$ and P-wave baryons are also discussed, we find that the leading contributions to the isospin amplitudes at $O(\mu^2)$ are from the transition between the P-wave baryons and the nucleon and the charge radius of the nucleon. The leading contribution from the P-wave baryons only affects the neutral pion production, and improve the agreement with data significantly. The transition between the resonance $P_{33}(1232)$ and the nucleon only gives an order $\mu^3$ corrections to $A^{(-)}$

E. coli elongation factor Tu bound to a GTP analogue displays an open conformation equivalent to the GDP-bound form

According to the traditional view, GTPases act as molecular switches, which cycle between distinct ‘on’ and ‘off’ conformations bound to GTP and GDP, respectively. Translation elongation factor EF-Tu is a GTPase essential for prokaryotic protein synthesis. In its GTP-bound form, EF-Tu delivers aminoacylated tRNAs to the ribosome as a ternary complex. GTP hydrolysis is thought to cause the release of EF-Tu from aminoacyl-tRNA and the ribosome due to a dramatic conformational change following Pi release. Here, the crystal structure of Escherichia coli EF-Tu in complex with a non-hydrolysable GTP analogue (GDPNP) has been determined. Remarkably, the overall conformation of EF-Tu·GDPNP displays the classical, open GDP-bound conformation. This is in accordance with an emerging view that the identity of the bound guanine nucleotide is not ‘locking’ the GTPase in a fixed conformation. Using a single molecule approach, the conformational dynamics of various ligand-bound forms of EF-Tu were probed in solution by fluorescence resonance energy transfer. The results suggest that EF-Tu, free in solution, may sample a wider set of conformations than the structurally well-defined GTP- and GDP-forms known from previous X-ray crystallographic studies. Only upon binding, as a ternary complex, to the mRNA programmed ribosome, is the well-known, closed GTP-bound conformation, observed

Research Notes : United States : Superoxide dismutase (SOD) in soybean

We are using vertical polyacrylamide gel electrophoresis (Davis, 1964) and a staining system modified after Beauchamp and Fridovich (1971) to study superoxide dismutase polymorphisms in the subgenus soja. This staining system generates superoxide radical; hence, it is specific for SOD activity. We re-solve up to 9 SOD bands in dry or germinating soybean cotyledons, and in leaves

Automatic radiographic quantification of hand osteoarthritis; accuracy and sensitivity to change in joint space width in a phantom and cadaver study

This is the final version of the article. Available from Springer Verlag via the DOI in this record.OBJECTIVE: To validate a newly developed quantification method that automatically detects and quantifies the joint space width (JSW) in hand radiographs. Repeatability, accuracy and sensitivity to changes in JSW were determined. The influence of joint location and joint shape on the measurements was tested. METHODS: A mechanical micrometer set-up was developed to define and adjust the true JSW in an acrylic phantom joint and in human cadaver-derived phalangeal joints. Radiographic measurements of the JSW were compared to the true JSW. Repeatability, systematic error (accuracy) and sensitivity (defined as the smallest detectable difference (SDD)) were determined. The influence of joint position on the JSW measurement was assessed by varying the location of the acrylic phantom on the X-ray detector with respect to the X-ray beam and the influence of joint shape was determined by using morphologically different human cadaver joints. RESULTS: The mean systematic error was 0.052 mm in the phantom joint and 0.210 mm in the cadaver experiment. In the phantom experiments, the repeatability was high (SDD = 0.028 mm), but differed slightly between joint locations (p = 0.046), and a change in JSW of 0.037 mm could be detected. Dependent of the joint shape in the cadaver hand, a change in JSW between 0.018 and 0.047 mm could be detected. CONCLUSIONS: The automatic quantification method is sensitive to small changes in JSW. Considering the published data of JSW decline in the normal and osteoarthritic population, the first signs of OA progression with this method can be detected within 1 or 2 years.This work was funded by the Dutch Arthritis Association (Reumafonds). The study sponsor had no involvement in study design, data collection, data analysis, or interpretation of the results