Molecular identification of three Arabidopsis thaliana mitochondrial dicarboxylate carrier isoforms: organ distribution, bacterial expression, reconstitution into liposomes and functional characterization

Screening of the Arabidopsis thaliana genome revealed three potential homologues of mammalian and yeast mitochondrial DICs (dicarboxylate carriers) designated as DIC1, DIC2 and DIC3, each belonging to the mitochondrial carrier protein family. DIC1 and DIC2 are broadly expressed at comparable levels in all the tissues investigated. DIC1-DIC3 have been reported previously as uncoupling proteins, but direct transport assays with recombinant and reconstituted DIC proteins clearly demonstrate that their substrate specificity is unique to plants, showing the combined characteristics of the DIC and oxaloacetate carrier in yeast. Indeed, the Arabidopsis DICs transported a wide range of dicarboxylic acids including malate, oxaloacetate and succinate as well as phosphate, sulfate and thiosulfate at high rates, whereas 2-oxoglutarate was revealed to be a very poor substrate. The role of these plant mitochondrial DICs is discussed with respect to other known mitochondrial carrier family members including uncoupling proteins. It is proposed that plant DICs constitute the membrane component of several metabolic processes including the malate-oxaloacetate shuttle, the most important redox connection between the mitochondria and the cytosol.     

Chloroplast nitrite uptake is enhanced in Arabidopsis PII mutants

In higher plants, the PII protein is a nuclear-encoded plastid protein that regulates the activity of a key enzyme of arginine biosynthesis. We have previously observed that Arabidopsis PII mutants are more sensitive to nitrite toxicity. Using intact chloroplasts isolated from Arabidopsis leaves and (15)N-labelled nitrite we show that a light-dependent nitrite uptake into chloroplasts is increased in PII knock-out mutants when compared to the wild-type. This leads to a higher incorporation of (15)N into ammonium and amino acids in the mutant chloroplasts. However, the uptake differences do not depend on GS/GOGAT activities. Our observations suggest that PII is involved in the regulation of nitrite uptake into higher plant chloroplasts.     

Modeling longitudinal spatial periodontal data: a spatially adaptive model with tools for specifying priors and checking fit

Summary .   Attachment loss (AL), the distance down a tooth's root that is no longer attached to surrounding bone by periodontal ligament, is a common measure of periodontal disease. In this article, we develop a spatiotemporal model to monitor the progression of AL. Our model is an extension of the conditionally autoregressive (CAR) prior, which spatially smooths estimates toward their neighbors. However, because AL often exhibits a burst of large values in space and time, we develop a nonstationary spatiotemporal CAR model that allows the degree of spatial and temporal smoothing to vary in different regions of the mouth. To do this, we assign each AL measurement site its own set of variance parameters and spatially smooth the variances with spatial priors. We propose a heuristic to measure the complexity of the site-specific variances, and use it to select priors that ensure parameters in the model are well identified. In data from a clinical trial, this model improves the fit compared to the usual dynamic CAR model for 90 of 99 patients' AL measurements.     

Stability and specificity of heterodimer formation for the coiled-coil neck regions of the motor proteins Kif3A and Kif3B: the role of unstructured oppositely charged regions.

We investigated the folding, stability, and specificity of dimerization of the neck regions of the kinesin-like proteins Kif3A (residues 356-416) and Kif3B (residues 351-411). We showed that the complementary charged regions found in the hinge regions (which directly follow the neck regions) of these proteins do not adopt any secondary structure in solution. We then explored the ability of the complementary charged regions to specify heterodimer formation for the neck region coiled-coils found in Kif3A and Kif3B. Redox experiments demonstrated that oppositely charged regions specified the formation of a heterodimeric coiled-coil. Denaturation studies with urea demonstrated that the negatively charged region of Kif3A dramatically destabilized its neck coiled-coil (urea1/2 value of 3.9 m compared with 6.7 m for the coiled-coil alone). By comparison, the placement of a positively charged region C-terminal to the neck coiled-coil of Kif3B had little effect on stability (urea1/2 value of 8.2 m compared with 8.8 m for the coiled-coil alone). The pairing of complementary charged regions leads to specific heterodimer formation where the stability of the heterodimeric neck coiled-coil with charged regions had similar stability (urea1/2 value of 7.8 m) to the most stable homodimer (Kif3B) with charged regions (urea1/2 value of 8.0 m) and dramatically more stable than the Kif3A homodimer with charged regions (urea1/2, value of 3.9 m). The heterodimeric coiled-coil with charged extensions has essentially the same stability as the heterodimeric coiled-coil on its own (urea1/2 values of 7.8 and 8.1 m, respectively) suggesting that specificity of heterodimerization is driven by non-specific attraction of the oppositely unstructured charged regions without affecting stability of the heterodimeric coiled-coil.     

Mycobacterial and nonbacterial pulmonary complications in hospitalized patients with human immunodeficiency virus infection: A prospective, cohort study

Background  

A prospective observational study was done to describe nonbacterial pulmonary complications in hospitalized patients with human immunodeficiency virus (HIV) infection.     

Physiological and chemical indicators for early and late stages of sepsis in conscious rats

Endotoxin shock is a major cause of death in patients with septicemia. Endotoxin induces nitric oxide (NO) production and causes tissue damage. In addition, the release of oxygen free radicals has also been observed in endotoxin shock and was found to be responsible for the occurrence of multiple organ failure. The purpose of the present study was to evaluate suitable indicators for early and late stages of endotoxin shock. The experiments were designed to induce endotoxin shock in conscious rats by means of anEscherichia coli lipopolysaccharide (LPS) injection. Arterial pressure (AP) and heart rate (HR) were continuously monitored for 72 h after LPS administration. The maximal decrease in AP and increase in HR and nitrate/nitrite level occurred at 9–12 h following LPS administration. The white blood cell (WBC) count had decreased at 3 h. Hydroxyl radical (methyl guanidine, MG) decreased rapidly after LPS administration. Plasma levels of blood urea nitrogen (BUN), creatinine (Cr), lactic dehydrogenase (LDH), creatine phosphokinase (CPK), and glutamic oxaloacetic transaminase increased before the rise of amylase. Our results suggest that changes in AP, HR, WBC, free radicals, and chemical substances (BUN, Cr) can possibly serve as approximate indicators for the early stage of endotoxin shock. Severe multiple organ damage may be caused by amylase release in the late stage of endotoxin shock.     

Determination of stereochemistry stability coefficients of amino acid side-chains in an amphipathic alpha-helix.

We describe here a systematic study to determine the effect on secondary structure of d-amino acid substitutions in the nonpolar face of an amphipathic alpha-helical peptide. The helix-destabilizing ability of 19 d-amino acid residues in an amphipathic alpha-helical model peptide was evaluated by reversed-phase HPLC and CD spectroscopy. l-Amino acid and d-amino acid residues show a wide range of helix-destabilizing effects relative to Gly, as evidenced in melting temperatures (DeltaTm) ranging from -8.5 degrees C to 30.5 degrees C for the l-amino acids and -9.5 degrees C to 9.0 degrees C for the d-amino acids. Helix stereochemistry stability coefficients defined as the difference in Tm values for the l- and d-amino acid substitutions [(DeltaTm' = TmL and TmD)] ranging from 1 degrees C to 34.5 degrees C. HPLC retention times [DeltatR(XL-XD)] also had values ranging from -0.52 to 7.31 min at pH 7.0. The helix-destabilizing ability of a specific d-amino acid is highly dependent on its side-chain, with no clear relationship to the helical propensity of its corresponding l-enantiomers. In both CD and reversed-phase HPLC studies, d-amino acids with beta-branched side-chains destabilize alpha-helical structure to the greatest extent. A series of helix stability coefficients was subsequently determined, which should prove valuable both for protein structure-activity studies and de novo design of novel biologically active peptides.