Predicting RNA secondary structures with pseudoknots by MCMC sampling

The most probable secondary structure of an RNA molecule, given the nucleotide sequence, can be computed efficiently if a stochastic context-free grammar (SCFG) is used as the prior distribution of the secondary structure. The structures of some RNA molecules contain so-called pseudoknots. Allowing all possible configurations of pseudoknots is not compatible with context-free grammar models and makes the search for an optimal secondary structure NP-complete. We suggest a probabilistic model for RNA secondary structures with pseudoknots and present a Markov-chain Monte-Carlo Method for sampling RNA structures according to their posterior distribution for a given sequence. We favor Bayesian sampling over optimization methods in this context, because it makes the uncertainty of RNA structure predictions assessable. We demonstrate the benefit of our method in examples with tmRNA and also with simulated data. McQFold, an implementation of our method, is freely available from http://www.cs.uni-frankfurt.de/~metzler/McQFold.     

Energy dissipation is an essential mechanism to sustain the viability of plants: The physiological limits of improved photosynthesis

In bright sunlight photosynthetic activity is limited by the enzymatic machinery of carbon dioxide assimilation. This supererogation of energy can be easily visualized by the significant increases of photosynthetic activity under high CO₂ conditions or other metabolic strategies which can increase the carbon flux from CO₂ to metabolic pools. However, even under optimal CO₂ conditions plants will provide much more NADPH + H⁺ and ATP that are required for the actual demand, yielding in a metabolic situation, in which no reducible NADP⁺ would be available. As a consequence, excited chlorophylls can activate oxygen to its singlet state or the photosynthetic electrons can be transferred to oxygen, producing highly active oxygen species such as the superoxide anion, hydroxyl radicals and hydrogen peroxide. All of them can initiate radical chain reactions which degrade proteins, pigments, lipids and nucleotides. Therefore, the plants have developed protection and repair mechanism to prevent photodamage and to maintain the physiological integrity of metabolic apparatus. The first protection wall is regulatory energy dissipation on the level of the photosynthetic primary reactions by the so-called non-photochemical quenching. This dissipative pathway is under the control of the proton gradient generated by the electron flow and the xanthophyll cycle. A second protection mechanism is the effective re-oxidation of the reduction equivalents by so-called “alternative electron cycling” which includes the water-water cycle, the photorespiration, the malate valve and the action of antioxidants. The third system of defence is the repair of damaged components. Therefore, plants do not suffer from energy shortage, but instead they have to invest in proteins and cellular components which protect the plants from potential damage by the supererogation of energy. Under this premise, our understanding and evaluation for certain energy dissipating processes such as non-photochemical quenching or photorespiration appear in a quite new perspective, especially when discussing strategies to improve the solar energy conversion into plant biomass.     

Autism and X-linked hypophosphatemia: A possible association?

We herein report the joint occurrence of an autistic disorder (AD) and X-linked hypophosphatemia. X-linked hypophosphatemia (XLH), an X-linked dominant disorder, is the most common of the inherited renal phosphate wasting disorders. Autism is a pervasive developmental disorder that occurs mainly due to genetic causes. In approximately 6-15% of cases, the autistic phenotype is a part of a broader genetic condition called syndromic autism.Therefore, reports of cases with the joint occurrence of a known genetic syndrome and a diagnosis of ASD by a child psychiatrist are relevant. A joint occurrence does not, however, mean that there is always a causal link between the genetic syndrome and the autistic behavioural phenotype. In this case, there are a number of arguments countering a causal link.     

Classification of subcellular location by comparative proteomic analysis of native and density-shifted lysosomes

One approach to the functional characterization of the lysosome lies in the use of proteomic methods to identify proteins in subcellular fractions enriched for this organelle. However, distinguishing between true lysosomal residents and proteins from other cofractionating organelles is challenging. To this end, we implemented a quantitative mass spectrometry approach based on the selective decrease in the buoyant density of liver lysosomes that occurs when animals are treated with Triton-WR1339. Liver lysosome-enriched preparations from control and treated rats were fractionated by isopycnic sucrose density gradient centrifugation. Tryptic peptides derived from gradient fractions were reacted with isobaric tag for relative and absolute quantitation eight-plex labeling reagents and analyzed by two-dimensional liquid chromatography matrix-assisted laser desorption ionization time-of-flight MS. Reporter ion intensities were used to generate relative protein distribution profiles across both types of gradients. A distribution index was calculated for each identified protein and used to determine a probability of lysosomal residence by quadratic discriminant analysis. This analysis suggests that several proteins assigned to the lysosome in other proteomics studies are not true lysosomal residents. Conversely, results support lysosomal residency for other proteins that are either not or only tentatively assigned to this location. The density shift for two proteins, Cu/Zn superoxide dismutase and ATP-binding cassette subfamily B (MDR/TAP) member 6, was corroborated by quantitative Western blotting. Additional balance sheet analyses on differential centrifugation fractions revealed that Cu/Zn superoxide dismutase is predominantly cytosolic with a secondary lysosomal localization whereas ATP-binding cassette subfamily B (MDR/TAP) member 6 is predominantly lysosomal. These results establish a quantitative mass spectrometric/subcellular fractionation approach for identification of lysosomal proteins and underscore the necessity of balance sheet analysis for localization studies.     

Parallel Exploitation of Diverse Host Nutrients Enhances Salmonella Virulence

Pathogen access to host nutrients in infected tissues is fundamental for pathogen growth and virulence, disease progression, and infection control. However, our understanding of this crucial process is still rather limited because of experimental and conceptual challenges. Here, we used proteomics, microbial genetics, competitive infections, and computational approaches to obtain a comprehensive overview of Salmonella nutrition and growth in a mouse typhoid fever model. The data revealed that Salmonella accessed an unexpectedly diverse set of at least 31 different host nutrients in infected tissues but the individual nutrients were available in only scarce amounts. Salmonella adapted to this situation by expressing versatile catabolic pathways to simultaneously exploit multiple host nutrients. A genome-scale computational model of Salmonella in vivo metabolism based on these data was fully consistent with independent large-scale experimental data on Salmonella enzyme quantities, and correctly predicted 92% of 738 reported experimental mutant virulence phenotypes, suggesting that our analysis provided a comprehensive overview of host nutrient supply, Salmonella metabolism, and Salmonella growth during infection. Comparison of metabolic networks of other pathogens suggested that complex host/pathogen nutritional interfaces are a common feature underlying many infectious diseases.     

Mapping the Binding Interface between an HIV-1 Inhibiting Intrabody and the Viral Protein Rev

HIV-1 Rev is the key protein in the nucleocytoplasmic export and expression of the late viral mRNAs. An important aspect for its function is its ability to multimerize on these mRNAs. We have recently identified a llama single-domain antibody (Nb₁₉₀) as the first inhibitor targeting the Rev multimerization function in cells. This nanobody is a potent intracellular antibody that efficiently inhibits HIV-1 viral production. In order to gain insight into the Nb₁₉₀-Rev interaction interface, we performed mutational and docking studies to map the interface between the nanobody paratope and the Rev epitope. Alanine mutants of the hyper-variable domains of Nb₁₉₀ and the Rev multimerization domains were evaluated in different assays measuring Nb₁₉₀-Rev interaction or viral production. Seven residues within Nb₁₉₀ and five Rev residues are demonstrated to be crucial for epitope recognition. These experimental data were used to perform docking experiments and map the Nb₁₉₀-Rev structural interface. This Nb₁₉₀-Rev interaction model can guide further studies of the Nb₁₉₀ effect on HIV-1 Rev function and could serve as starting point for the rational development of smaller entities binding to the Nb₁₉₀ epitope, aimed at interfering with protein-protein interactions of the Rev N-terminal domain.     

Pleiotropic consequences of mutations towards antibiotic-hypersensitivity in Serratia marcescens

Various mutants (oxa ^s ) were isolated from Serratia marcescens SM-6 by selecting for hypersensitivity towards oxacillin. All mutants found are highly pleiotropic and able to yield spontaneous revertants which behave like the wild-type. Mutant W 1421 mostly studied shows the following phenotypic properties not found in the wild-type: (1) The growth is hypersensitive to various antibiotics, detergents and dyes which differ remarkably in their chemical structure and antibacterial action-mechanism, (2) the cells can be easily solubilized by 0.05% Sodium-dodecylsulfate, (3) the cells allow the adsorption of the roughmutant specific Salmonella phage 6SR, (4) strong cellular binding of crystal violet, (5) agglutination of the cells in 0.3% auramin solution and (6) reduced formation of red pigment. Strain W 1421 is assumed to be a lipopolysaccharide-defective mutant. The outer membrane of mutant W 1421 analyzed by Sodiumdodecylsulfate-polyacrylamide gel electrophoresis possesses a single protein less than that of the wild-type. Mutant W 1421 is further characterized by its low exolipase activity; exoprotease and exonuclease activities are as in the wild-type. This specific exoenzyme deficiency can be overcome either by backmutation to oxacillin-resistance or by growing mutant W 1421 in a medium supplemented with certain non-metabolizable polysaccharides, e.g. glycogen or pectin B. Both polysaccharides increase the exolipase activity of the wild-type too.List of Abbreviations amp ampicillin - LPS lipopolysaccharide - MIC minimal inhibitory concentration - NB nutrient broth - oxa oxacillin - str streptomycin - TBY tryptone broth with yeast extract - SDS sodium-dodecylsulfate - OD optical density This paper is dedicated to Prof. Dr. R. W. Kaplan, University of Frankfurt/M., on the occasion of his 65th birthday     

Combined molecular and morphological data provide insights into the evolution and classification of Chilocorini ladybirds (Coleoptera: Coccinellidae)

Ladybirds of the cosmopolitan tribe Chilocorini prey mainly on coccids and include several important biocontrol agents. The phylogenetic relationships of Chilocorini are poorly known. In this paper, we provide a phylogenetic reconstruction of Chilocorini containing all 27 genera based on five molecular markers and 86 adult morphological characters. Morphological character states were mapped on the combined data tree from Bayesian inference to analyse morphological traits of each genus. Sixteen morphological characters were selected to reconstruct the ancestral states using maximum parsimony and maximum likelihood methods. Divergence times were estimated based on the relaxed molecular clock approach. Our results indicate that Chilocorini, excluding Chilocorellus Miyatake, is monophyletic and closely related to Plotinini. The crown group Chilocorini was estimated to date back to the Middle Cretaceous. Anisorcus Crotch, Egius Mulsant, Phaenochilus Weise and Simmondsius Ahmad & Ghani are synonymized here with Chilocorus Leach ( syn.n. ). The genus Chilocorellus is excluded from Chilocorini. The split of current genera was estimated to have occurred during the Middle Paleogene to Late Paleogene.     

Cyclooxygenase and 5-lipoxygenase inhibitory activity of 2,6 di-t-butylphenols linked by a sulfur atom to 1,3,4-thiadiazoles and 1,3,4-oxadiazoles

The preparation of a series of 1,3,4-thiadiazoles and 1,3,4-oxadizoles linked by a thioether to 2,6-di-t-butylphenol and the inhibition of cyclooxygenase (CO) and 5-lipoxygenase (5-LO) by these compounds is dicussed.