Phospho-ubiquitin-PARK2 complex as a marker for mitophagy defects

The E3 ubiquitin ligase PARK2 and the mitochondrial protein kinase PINK1 are required for the initiation of mitochondrial damage-induced mitophagy. Together, PARK2 and PINK1 generate a phospho-ubiquitin signal on outer mitochondrial membrane proteins that triggers recruitment of the autophagy machinery. This paper describes the detection of a defined 500-kDa phospho-ubiquitin-rich PARK2 complex that accumulates on mitochondria upon treatment with the membrane uncoupler CCCP. Formation of this complex is dependent on the presence of PINK1 and is absent in mutant forms of PARK2, whereby mitophagy is also arrested. These results signify a functional signaling complex that is essential for the progression of mitophagy. The visualization of the PARK2 signaling complex represents a novel marker for this critical step in mitophagy and can be used to monitor mitophagy progression in PARK2 mutants and to uncover additional upstream factors required for PARK2-mediated mitophagy signaling.     

Identification of Cell Cycle Dependent Interaction Partners of the Septins by Quantitative Mass Spectrometry

The septins are a conserved family of GTP-binding proteins that, in the baker''s yeast, assemble into a highly ordered array of filaments at the mother bud neck. These filaments undergo significant structural rearrangements during the cell cycle. We aimed at identifying key components that are involved in or regulate the transitions of the septins. By combining cell synchronization and quantitative affinity-purification mass-spectrometry, we performed a screen for specific interaction partners of the septins at three distinct stages of the cell cycle. A total of 83 interaction partners of the septins were assigned. Surprisingly, we detected DNA-interacting/nuclear proteins and proteins involved in ribosome biogenesis and protein synthesis predominantly present in alpha-factor arrested that do not display an assembled septin structure. Furthermore, two distinct sets of regulatory proteins that are specific for cells at S-phase with a stable septin collar or at mitosis with split septin rings were identified.Complementary methods like SPLIFF and immunoprecipitation allowed us to more exactly define the spatial and temporal characteristics of selected hits of the AP-MS screen.     

Oat mesophyll protoplasts: their response to various feeder cultures

Oat (Avena sativa L.) mesophyll protoplasts were recently demonstrated to be capable of dedifferentiation, repeated divisions, and colony formation. Since the development of oat mesophyll protoplasts is decisively influenced by the nature of the used feeder culture (species, variety and concentration), we conducted a systematic study of this parameter. Generally, graminaceous feeders promoted protoplast proliferation, while dicot species repressed protoplast divisions. The beneficial effect of those feeders that promote divisions was counterbalanced by a factor that causes necrosis. The correct balance between promotion of divisions or necrosis depended on the nature of the feeder and its plating density.     

Reconstruction of Cellular Signal Transduction Networks Using Perturbation Assays and Linear Programming

Perturbation experiments for example using RNA interference (RNAi) offer an attractive way to elucidate gene function in a high throughput fashion. The placement of hit genes in their functional context and the inference of underlying networks from such data, however, are challenging tasks. One of the problems in network inference is the exponential number of possible network topologies for a given number of genes. Here, we introduce a novel mathematical approach to address this question. We formulate network inference as a linear optimization problem, which can be solved efficiently even for large-scale systems. We use simulated data to evaluate our approach, and show improved performance in particular on larger networks over state-of-the art methods. We achieve increased sensitivity and specificity, as well as a significant reduction in computing time. Furthermore, we show superior performance on noisy data. We then apply our approach to study the intracellular signaling of human primary nave CD4⁺ T-cells, as well as ErbB signaling in trastuzumab resistant breast cancer cells. In both cases, our approach recovers known interactions and points to additional relevant processes. In ErbB signaling, our results predict an important role of negative and positive feedback in controlling the cell cycle progression.     

Recent advances in imine reductase-catalyzed reactions

Imine reductases are nicotinamide-dependent enzymes that catalyze the asymmetric reduction of various imines to the corresponding amine products. Owing to the increasing roles of chiral amines and heterocyclic compounds as intermediates for pharmaceuticals, the demand for novel selective synthesis strategies is vitally important. Recent studies have demonstrated the discovery and structural characterization of a number of stereoselective imine reductase enzymes. Here, we highlight recent progress in applying imine reductases for the formation of chiral amines and heterocycles. It particularly focuses on the utilization of imine reductases in reductive aminations of aldehydes and ketones with various amine nucleophiles, one of the most powerful reactions in the synthesis of chiral amines. Second, we report on the synthesis of saturated substituted N-heterocycles by combining them with further biocatalysts, such as carboxylic acid reductases, oxidases or transaminases. Finally, we summarize the latest applications of imine reductases in the promiscuous asymmetric hydrogenation of a highly reactive carbonyl compound and the engineering of the cofactor specificity from NADPH to NADH.     

Yersinia outer protein YopE affects the actin cytoskeleton in Dictyostelium discoideum through targeting of multiple Rho family GTPases

Background  

All human pathogenic Yersinia species share a virulence-associated type III secretion system that translocates Yersinia effector proteins into host cells to counteract infection-induced signaling responses and prevent phagocytosis. Dictyostelium discoideum has been recently used to study the effects of bacterial virulence factors produced by internalized pathogens. In this study we explored the potential of Dictyostelium as model organism for analyzing the effects of ectopically expressed Yersinia outer proteins (Yops).     

Core area analysis at semi-deciduous forest islands in the Comoé National Park,NE Ivory Coast

For the protection of forest-interior species in both natural forest islands and anthropogenic forest fragments knowledge on the size of forest-core areas is a central issue. In an intact mosaic of semi-deciduous forests and savanna in the Comoé National Park 31 forest islands were selected (2.1–146.1 ha). Values for the depth-of-edge influence (DEI) of the study area recently published range from 0 m up to nearly 150 m. Thus, core-area analysis was carried out for this range in 5 m steps. For a DEI of 55 m—e.g. computed for tree-species composition of large trees—half of the total forest area can be considered as core area, but only 9 of the studied forest islands still contained a relative core area (rCA) of more than 50%. From non-linear regression it was estimated that for a DEI of 55 m an rCA of 50% can be expected for forest islands with a size of 36.6 ± 7.6 ha. This value increased exponentially with increasing DEI. The GIS-based core-area analysis presented in this paper proved to be suitable to give a well interpretable overview on rCA with respect to varying DEI, and we recommend to incorporate this type of analysis in existing GIS-tools. As the presented study is the first sound core area analysis at forest islands in West Africa, data contribute to a better understanding of this field of ecology that is of high relevance for planners and decision makers to protect biodiversity.