Oenothera mitochondrial orf454, a gene involved in cytochrome c biogenesis corresponds to orf169 and orf322 of Marchantia

We have characterized a mitochondrial gene in Oenothera, designated orf454, capable of encoding a component of the cytochrome c biogenesis system. This open reading frame is interrupted by an intron of 941 nucleotides showing high similarity to a group II intron residing in the rpl2 gene. RNA editing, which is observed at 18 cytidine positions within the orf454 reading frame, improves the similarity to protein-coding sequences in bacteria and higher plants and removes the last 16 amino acids. orf454 also shows high sequence similarity to two overlapping reading frames (orf169 and orf322) of Marchantia mitochondria. These ORFs belong to an operon-like cluster of genes in the liverwort that is not conserved in Oenothera mitochondria. However, in bacteria these reading frames are organized like the Marchantia gene cluster. It has been shown by genetical analysis in Rhodobacter capsulatus that these genes are essential for cytochrome c biogenesis. Genes of bacterial operons — ccl1 in Rhodobacter and yejR and nrfE in Escherichia coli — show high sequence similarity to the mitochondrial reading frames orf577 and orf454 of Oenothera. orf454, which we describe here, is homologous to the C-terminal region of these bacterial genes, while the previously described orf577 is homologous to the N-terminal region.     

Porcine arterivirus attachment to the macrophage-specific receptor sialoadhesin is dependent on the sialic acid-binding activity of the N-terminal immunoglobulin domain of sialoadhesin

The sialic acid-binding lectin sialoadhesin (Sn) is a macrophage-restricted receptor for porcine reproductive and respiratory syndrome virus (PRRSV). To investigate the importance of pSn sialic acid-binding activity for PRRSV infection, an R₁₁₆-to-E mutation was introduced in the predicted sialic acid-binding domain of pSn, resulting in a mutant, pSn^RE, that could not bind sialic acids. PSn, but not pSn^RE, allowed PRRSV binding and internalization. These data show that the sialic acid-binding activity of pSn is essential for PRRSV attachment to pSn and thus identifies the variable, N-terminal domain of Sn as a PRRSV binding domain.     

Metabolism and foraging strategies of mid‐latitude mesozooplankton during cyanobacterial blooms as revealed by fatty acids,amino acids,and their stable carbon isotopes

Increasing sea surface temperatures (SST) and blooms of lipid‐poor, filamentous cyanobacteria can change mesozooplankton metabolism and foraging strategies in marine systems. Lipid shortage and imbalanced diet may challenge the build‐up of energy pools of lipids and proteins, and access to essential fatty acids (FAs) and amino acids (AAs) by copepods. The impact of cyanobacterial blooms on individual energy pools was assessed for key species temperate Temora longicornis and boreal Pseudo‐/Paracalanus spp. that dominated field mesozooplankton communities isolated by seasonal stratification in the central Baltic Sea during the hot and the cold summer. We looked at (a) total lipid and protein levels, (b) FA trophic markers and AA composition, and (c) compound‐specific stable carbon isotopes (δ¹³C) in bulk mesozooplankton and in a subset of parameters in particulate organic matter. Despite lipid‐poor cyanobacterial blooms, the key species were largely able to cover both energy pools, yet a tendency of lipid reduction was observed in surface animals. Omni‐ and carnivory feeding modes, FA trophic makers, and δ¹³C patterns in essential compounds emphasized that cyanobacterial FAs and AAs have been incorporated into mesozooplankton mainly via feeding on mixo‐ and heterotrophic (dino‐) flagellates and detrital complexes during summer. Foraging for essential highly unsaturated FAs from (dino‐) flagellates may have caused night migration of Pseudo‐/Paracalanus spp. from the deep subhalocline waters into the upper waters. Only in the hot summer (SST>19.0°C) was T. longicornis submerged in the colder subthermocline water (~4°C). Thus, the continuous warming trend and simultaneous feeding can eventually lead to competition on the preferred diet by key copepod species below the thermocline in stratified systems. A comparison of δ¹³C patterns of essential AAs in surface mesozooplankton across sub‐basins of low and high cyanobacterial biomasses revealed the potential of δ¹³C‐AA isoscapes for studies of commercial fish feeding trails across the Baltic Sea food webs.     

Sustainable control of mosquito larvae in the field by the combined actions of the biological insecticide Bti and natural competitors

Integrated management of mosquitoes is becoming increasingly important, particularly in relation to avoiding recolonization of ponds after larvicide treatment. We conducted for the first time field experiments that involved exposing natural populations of the mosquito species Culex pipiens to: a) application of the biological insecticide Bacillus thuringiensis israelensis (Bti), b) the introduction of natural competitors (a crustacean community composed mainly of Daphnia spp.), or c) a combined treatment that involved both introduction of a crustacean community and the application of Bti. The treatment that involved only the introduction of crustaceans had no significant effect on mosquito larval populations, while treatment with Bti alone caused only a significant reduction in the abundance of mosquito larvae in the short‐term (within 3–10 days after treatment). In contrast, the combined treatment rapidly reduced the abundance of mosquito larvae, which remained low throughout the entire observation period of 28 days. Growth of the introduced crustacean communities was favored by the immediate reduction in the abundance of mosquito larvae following Bti administration, thus preventing recolonization of ponds by mosquito larvae at the late period (days 14–28 after treatment). Both competition and the temporal order of establishment of different species are hence important mechanisms for efficient and sustainable mosquito control.     

Automated maintenance of embryonic stem cell cultures

Embryonic stem cell (ESC) technology provides attractive perspectives for generating unlimited numbers of somatic cells for disease modeling and compound screening. A key prerequisite for these industrial applications are standardized and automated systems suitable for stem cell processing. Here we demonstrate that mouse and human ESC propagated by automated culture maintain their mean specific growth rates, their capacity for multi-germlayer differentiation, and the expression of the pluripotency-associated markers SSEA-1/Oct-4 and Tra-1-60/Tra-1-81/Oct-4, respectively. The feasibility of ESC culture automation may greatly facilitate the use of this versatile cell source for a variety of biomedical applications.     

Human periosteum-derived progenitor cells express distinct chemokine receptors and migrate upon stimulation with CCL2, CCL25, CXCL8, CXCL12, and CXCL13

For bone repair, transplantation of periosteal progenitor cells (PCs), which had been amplified within supportive scaffolds, is applied clinically. More innovative bone tissue engineering approaches focus on the in situ recruitment of stem and progenitor cells to defective sites and their subsequent use for guided tissue repair. Chemokines are known to induce the directed migration of bone marrow CD34(-) mesenchymal stem cells (MSCs). The aim of our study was to determine the chemokine receptor expression profile of human CD34(-) PCs and to demonstrate that these cells migrate upon stimulation with selected chemokines. PCs were isolated from periosteum of the mastoid bone and displayed a homogenous cell population presenting an MSC-related cell-surface antigen profile (ALCAM(+), SH2(+), SH3(+), CD14(-), CD34(-), CD44(+), CD45(-), CD90(+)). The expression profile of chemokine receptors was determined by real-time PCR and immunohistochemistry. Both methods consistently demonstrated that PCs express receptors of all four chemokine subfamilies CC, CXC, CX(3)C, and C. Migration of PCs and a dose-dependent migratory effect of the chemokines CCL2 (MCP1), CCL25 (TECK), CXCL8 (IL8), CXCL12 (SDF1alpha), and CXCL13 (BCA1), but not CCL22 (MDC) were demonstrated using a 96-multiwell chemotaxis assay. In conclusion, for the first time, here we report that human PCs express chemokine receptors, present their profile, and demonstrate a dose-dependent migratory effect of distinct chemokines on these cells. These results are promising towards in situ bone repair therapies based on guiding PCs to bone defects, and encourage further in vivo studies.     

Assessment of three Resistance-Nodulation-Cell Division drug efflux transporters of Burkholderia cenocepacia in intrinsic antibiotic resistance

Background  

Burkholderia cenocepacia are opportunistic Gram-negative bacteria that can cause chronic pulmonary infections in patients with cystic fibrosis. These bacteria demonstrate a high-level of intrinsic antibiotic resistance to most clinically useful antibiotics complicating treatment. We previously identified 14 genes encoding putative Resistance-Nodulation-Cell Division (RND) efflux pumps in the genome of B. cenocepacia J2315, but the contribution of these pumps to the intrinsic drug resistance of this bacterium remains unclear.     

Structures of human MAP kinase kinase 1 (MEK1) and MEK2 describe novel noncompetitive kinase inhibition

MEK1 and MEK2 are closely related, dual-specificity tyrosine/threonine protein kinases found in the Ras/Raf/MEK/ERK mitogen-activated protein kinase (MAPK) signaling pathway. Approximately 30% of all human cancers have a constitutively activated MAPK pathway, and constitutive activation of MEK1 results in cellular transformation. Here we present the X-ray structures of human MEK1 and MEK2, each determined as a ternary complex with MgATP and an inhibitor to a resolution of 2.4 A and 3.2 A, respectively. The structures reveal that MEK1 and MEK2 each have a unique inhibitor-binding pocket adjacent to the MgATP-binding site. The presence of the potent inhibitor induces several conformational changes in the unphosphorylated MEK1 and MEK2 enzymes that lock them into a closed but catalytically inactive species. Thus, the structures reported here reveal a novel, noncompetitive mechanism for protein kinase inhibition.     

Structural investigations on betacyanin pigments by LC NMR and 2D NMR spectroscopy

Four betacyanin pigments were analysed by LC NMR and subjected to extensive NMR characterisation after isolation. Previously, low pH values were applied for NMR investigations of betalains resulting in rapid degradation of the purified substances thus preventing extensive NMR studies. Consequently, up to now only one single (13)C NMR spectrum of a betalain pigment, namely that of neobetanin (=14,15-dehydrobetanin), was available. Because of its sufficient stability under highly acidic conditions otherwise detrimental for betacyanins, this pigment remained an exemption. Since betalains are most stable in the pH range of 5-7, a new solvent system has been developed allowing improved data acquisition through improved pigment stability at near neutral pH. Thus, not only (1)H, but for the first time also partial (13)C data of betanin, isobetanin, phyllocactin and hylocerenin isolated from red-purple pitaya [Hylocereus polyrhizus (Weber) Britton & Rose, Cactaceae] could be indirectly obtained by gHSQC- and gHMQC-NMR experiments.