Mononuclear and dinuclear bromo bridged iridium(I) complexes with N-allyl substituted imidazolin-2-ylidene ligands

The reaction of 1-methyl-3-(2-propenyl)imidazolium bromide (1) or 1,3-bis(2-propenyl)-imidazolium bromide (2) with [Ir(μ-OMe)(cod)]₂ afforded the five coordinated iridium(I) carbene complexes [IrBr(L)(cod)] (3) (L=1-methyl-3-(2-propenyl)imidazolin-2-ylidene) and (4) (L=1,3-bis(2-propenyl)imidazolin-2-ylidene). The reaction proceeds via an in situ deprotonation of the imidazolium salt. Molecular structure determinations on 3 and 4 confirmed the coordination of the carbene ligands via the carbene carbon atom and one allyl group in both complexes. Treatment of complex 3 with an excess of AgBF₄ gave the dinuclear bromo bridged complex [(Ir(μ-Br)(L)(cod)]₂BF₄ (5) (L=1-methyl-3-(2-propenyl)imidazolin-2-ylidene). The reaction of complex 4 with an excess of AgBF₄ led to the mononuclear complex [Ir(L)(cod)]BF₄ (6) (L=1,3-bis(2-propenyl)imidazolin-2-ylidene) where both N-allyl substituents are coordinated to the iridium(I) center.     

Intracellular localization of RORalpha is isoform and cell line-dependent

The retinoid-related orphan receptor alpha (RORalpha) belongs to the nuclear receptor superfamily and comprises four isoforms generated by different promotor usage and alternative splicing. To better understand its function, the subcellular distribution of RORalpha was investigated. We could show that subcellular distribution of RORalpha is cell line and isoform-dependent. Isoform specific differences were mediated by the A/B domains which with the exception of RORalpha1 contain a signal that mediates cytoplasmic localization. The lack of this signal in RORalpha1 results in a complete nuclear localization and prevents cell membrane association observed for RORalpha2, 3, and 4. The region responsible for membrane association was identified as the C-terminal alpha-helix 12. Furthermore, the hinge region/ligand binding domain mediates nuclear localization. Our results show that isoform specific activity of RORalpha is not only regulated by different expression and DNA binding affinities but also by different subcellular distribution. Different access to the nucleus reveals an important mechanism regulating the activity of this constitutively active nuclear receptor.     

Mutations in PRP43 that uncouple RNA-dependent NTPase activity and pre-mRNA splicing function

Saccharomyces cerevisiae Prp43 is a DEAH-box RNA-dependent ATPase that catalyzes the release of excised lariat intron from the mRNA spliceosome. Previous studies identified mutations in Prp43 motifs I, II, and VI that were lethal in vivo and ablated ATP hydrolysis in vitro. Such Prp43 mutants exerted dominant-negative growth phenotypes when expressed in wild type cells and blocked intron release in vitro when added to yeast splicing extracts. Here, we assessed the effects of alanine and conservative substitutions at conserved residues in motifs Ia ((146)TQPRRVAA(153)), IV ((307)LLFLTG(312)), and V ((376)TNIAETSLT(384)) and thereby identified Arg150 (motif Ia), Phe309 (motif IV), Thr376, Leu383, and Thr384 (motif V) as being important for Prp43 function in vivo. Motif V mutations T376V, T384A, and T384V were lethal and dominant negative in vivo, and the mutant proteins inhibited lariat release in vitro. The T384A and T384V proteins were proficient for ATP hydrolysis, suggesting that ATPase activity is necessary, but not sufficient, for Prp43 function. We report that Prp43 hydrolyzes all common NTPs and dNTPs and unwinds short 5'/3' tailed RNA/DNA duplexes in an ATP-dependent fashion. Optimal ATP hydrolysis requires an RNA cofactor of >or=20 nt. Prp43 is largely indifferent to mutations in its C-terminal segment, which is conserved in the DEAH-box splicing factors Prp2, Prp16, and Prp22.     

Characterization of Bacillus strains from apple and pear trees in South Africa antagonistic to Erwinia amylovora

In order to find reasons for the absence of fire blight in most countries of the Southern hemisphere, bark samples from apple and pear trees in orchards of the Western Cape region in South Africa were extracted for bacteria which could be antagonistic to Erwinia amylovora. Screening was done in the late growth season and mainly Gram-positive bacteria were isolated. Approximately half of them produced growth inhibition zones on a lawn of E. amylovora. Most isolates were classified as Bacillus megaterium by microbiological assays and in API 50 test systems. They were visualized in the light microscope as non-motile large rods. These strains may not be responsible for the absence of fire blight in orchards, but they may indicate unfavourable climatic conditions for Gram-negative bacteria including E. amylovora. They may reduce the ability of E. amylovora to establish fire blight and could also be useful for application in biological disease control.     

<Emphasis Type="Italic">Puf</Emphasis> operon sequences and inferred structures of light-harvesting complexes of three closely related <Emphasis Type="Italic">Chromatiaceae</Emphasis> exhibiting different absorption characteristics

Whole cells of the purple sulfur bacterium strain 970 exhibit an unusual absorption peak at 963 nm. Its closest relatives, Thiorhodovibrio (Trv.) winogradskyi DSM6702^T and strain 06511 display a bacteriochlorophyll (BChl) a absorption peak at 867 nm that is characteristic for most light-harvesting complexes 1 (LHC1) of proteobacteria. The puf operons encoding the LHC1 and reaction center proteins were amplified, cloned, and sequenced, and for the Trv. winogradskyi, strains show the common pufBALMC gene arrangement, whereas strain 970 contains a second pufBA copy downstream of pufC. Only pufB ₁ A ₁ is transcribed, and the corresponding mRNA fragment had an increased stability. Alignments of the deduced protein sequences showed that the LHC1 polypeptides are closely related to those of Thermochromatium (Tch.) tepidum. A deletion between αHis⁰ and αTrp⁺¹¹, thought to be responsible for the redshifted Q _y absorption in Tch. tepidum, was also detected in strain 970 and Trv. winogradskyi, whereas αLys⁺¹² is replaced by histidine only in strain 970. Based on our structural modeling, the side chain of this αHis is predicted to be in close proximity to the BChl a, suggesting that it exerts a modulating effect on the spectral properties of the highly unusual LHC1 complex of strain 970.     

Mutations in SRCAP, encoding SNF2-related CREBBP activator protein, cause Floating-Harbor syndrome

Floating-Harbor syndrome (FHS) is a rare condition characterized by short stature, delayed osseous maturation, expressive-language deficits, and a distinctive facial appearance. Occurrence is generally sporadic, although parent-to-child transmission has been reported on occasion. Employing whole-exome sequencing, we identified heterozygous truncating mutations in SRCAP in five unrelated individuals with sporadic FHS. Sanger sequencing identified mutations in SRCAP in eight more affected persons. Mutations were de novo in all six instances in which parental DNA was available. SRCAP is an SNF2-related chromatin-remodeling factor that serves as a coactivator for CREB-binding protein (CREBBP, better known as CBP, the major cause of Rubinstein-Taybi syndrome [RTS]). Five SRCAP mutations, two of which are recurrent, were identified; all are tightly clustered within a small (111 codon) region of the final exon. These mutations are predicted to abolish three C-terminal AT-hook DNA-binding motifs while leaving the CBP-binding and ATPase domains intact. Our findings show that SRCAP mutations are the major cause of FHS and offer an explanation for the clinical overlap between FHS and RTS.     

Influence of air pressure, humidity, solar radiation, temperature, and wind speed on ambulatory visits due to chronic obstructive pulmonary disease in Bavaria, Germany

Chronic obstructive pulmonary disease (COPD) is one of the most important causes of morbidity and mortality in the world. The disease is often aggravated by periods of increased symptoms requiring medical attention. Among the possible triggers for these exacerbations, meteorological factors are under consideration. The objective of this study was to assess the influence of various meteorological factors on the health status of patients with COPD. For this purpose, the daily number of ambulatory care visits due to COPD was analysed in Bavaria, Germany, for the years 2006 and 2007. The meteorological factors were provided by the model at the European Centre for Medium Range Weather Forecast (ECMWF). For the multivariate analysis, a generalised linear model was used. In Bavaria, an increase of 1% of daily consultations (about 103 visits per day) was found to be associated with a change of 0.72?K temperature, 209.55 of log air surface pressure in Pa, and a decrease of 1% of daily consultations with 1,453,763 Ws m² of solar radiation. There also seem to be regional differences between north and south Bavaria; for instance, the effect of wind speed and specific humidity with a lag of 1?day were only significant in the north. This study could contribute to a tool for the prevention of exacerbations. It also serves as a model for the further evaluation of the impact of meteorological factors on health, and could easily be applied to other diseases or other regions.     

Comparative analysis of Neph gene expression in mouse and chicken development

Neph proteins are evolutionarily conserved members of the immunoglobulin superfamily of adhesion proteins and regulate morphogenesis and patterning of different tissues. They share a common protein structure consisting of extracellular immunoglobulin-like domains, a transmembrane region, and a carboxyl terminal cytoplasmic tail required for signaling. Neph orthologs have been widely characterized in invertebrates where they mediate such diverse processes as neural development, synaptogenesis, or myoblast fusion. Vertebrate Neph proteins have been described first at the glomerular filtration barrier of the kidney. Recently, there has been accumulating evidence suggesting a function of Neph proteins also outside the kidney. Here we demonstrate that Neph1, Neph2, and Neph3 are expressed differentially in various tissues during ontogenesis in mouse and chicken. Neph1 and Neph2 were found to be amply expressed in the central nervous system while Neph3 expression remained localized to the cerebellum anlage and the spinal cord. Outside the nervous system, Neph mRNAs were also differentially expressed in branchial arches, somites, heart, lung bud, and apical ectodermal ridge. Our findings support the concept that vertebrate Neph proteins, similarly to their Drosophila and C. elegans orthologs, provide guidance cues for cell recognition and tissue patterning in various organs which may open interesting perspectives for future research on Neph1-3 controlled morphogenesis.     

Serotonin signaling is required for Wnt-dependent GRP specification and leftward flow in Xenopus

In vertebrates, most inner organs are asymmetrically arranged with respect to the main body axis [1]. Symmetry breakage in fish, amphibian, and mammalian embryos depends on cilia-driven leftward flow of extracellular fluid during neurulation [2-5]. Flow induces the asymmetric nodal cascade that governs asymmetric organ morphogenesis and placement [1, 6, 7]. In the frog Xenopus, an alternative laterality-generating mechanism involving asymmetric localization of serotonin at the 32-cell stage has been proposed [8]. However, no functional linkage between this early localization and flow at neurula stage has emerged. Here, we report that serotonin signaling is required for specification of the superficial mesoderm (SM), which gives rise to the ciliated gastrocoel roof plate (GRP) where flow occurs [5, 9]. Flow and asymmetry were lost in embryos in which serotonin signaling was downregulated. Serotonin, which we found uniformly distributed along the main body axes in the early embryo, was required for Wnt signaling, which provides the instructive signal to specify the GRP. Importantly, serotonin was required for Wnt-induced double-axis formation as well. Our data confirm flow as primary mechanism of symmetry breakage and suggest a general role of serotonin as competence factor for Wnt signaling during axis formation in Xenopus.     

Impact of DAG stimulation on mineral synthesis, mineral structure and osteogenic differentiation of human cord blood stem cells

It remains unexplored in what way osteogenic stimulation with dexamethasone, ascorbic acid and β-glycerol phosphate (DAG) influences the process of mineralization, the composition and structure of the assembled mineral. Therefore, we analyzed and characterized biomineralization in DAG-stimulated and unstimulated 3D human unrestricted somatic stem cell (USSC) cultures. Microspheres were analyzed by histological staining, scanning electron microscopy (SEM), semi-quantitative energy-dispersive X-ray spectroscopy (EDX), quantitative wavelength-dispersive X-ray spectroscopy (WDX), transmission electron microscopy (TEM), selected area electron diffraction (SAED) and Raman spectroscopy.Mineral material was detected by SEM and histological staining in both groups, and showed structural differences. DAG influenced the differentiation of USSCs and the formation, structure and composition of the assembled mineral. SEM showed that cells of the + DAG spheres exhibited morphological signs of osteoblast-like cells. EDX and WDX confirmed a Ca-P mineral in both groups. Overall, the mineral material found showed structural similarities to the mineral substance of bony material.