Cytosolic Ras supports eye development in Drosophila

Ras proteins associate with cellular membranes as a consequence of a series of posttranslational modifications of a C-terminal CAAX sequence that include prenylation and are thought to be required for biological activity. In Drosophila melanogaster, Ras1 is required for eye development. We found that Drosophila Ras1 is inefficiently prenylated as a consequence of a lysine in the A(1) position of its CAAX sequence such that a significant pool remains soluble in the cytosol. We used mosaic analysis with a repressible cell marker (MARCM) to assess if various Ras1 transgenes could restore photoreceptor fate to eye disc cells that are null for Ras1. Surprisingly, we found that whereas Ras1 with an enhanced efficiency of membrane targeting could not rescue the Ras1 null phenotype, Ras1 that was not at all membrane targeted by virtue of a mutation of the CAAX cysteine was able to fully rescue eye development. In addition, constitutively active Ras1(12V,C186S) not targeted to membranes produced a hypermorphic phenotype and stimulated mitogen-activated protein kinase (MAPK) signaling in S2 cells. We conclude that the membrane association of Drosophila Ras1 is not required for eye development.     

Association between Type 2 Diabetes Loci and Measures of Fatness

Background

Type 2 diabetes (T2D) is a metabolic disorder characterized by disturbances of carbohydrate, fat and protein metabolism and insulin resistance. The majority of T2D patients are obese and obesity by itself may be a cause of insulin resistance. Our aim was to evaluate whether the recently identified T2D risk alleles are associated with human measures of fatness as characterized with Dual Energy X-ray Absorptiometry (DEXA).

Methodology/Principal Findings

Genotypes and phenotypes of approximately 3,000 participants from cross-sectional ERF study were analyzed. Nine single nucleotide polymorphisms (SNPs) in CDKN2AB, CDKAL1, FTO, HHEX, IGF2BP2, KCNJ11, PPARG, SLC30A8 and TCF7L2 were genotyped. We used linear regression to study association between individual SNPs and the combined allelic risk score with body mass index (BMI), fat mass index (FMI), fat percentage (FAT), waist circumference (WC) and waist to hip ratio (WHR). Significant association was observed between rs8050136 (FTO) and BMI (p = 0.003), FMI (p = 0.007) and WC (p = 0.03); fat percentage was borderline significant (p = 0.053). No other SNPs alone or combined in a risk score demonstrated significant association to the measures of fatness.

Conclusions/Significance

From the recently identified T2D risk variants only the risk variant of the FTO gene (rs8050136) showed statistically significant association with BMI, FMI, and WC.     

The major outer membrane protein OmpU of Vibrio splendidus contributes to host antimicrobial peptide resistance and is required for virulence in the oyster Crassostrea gigas

Vibrio splendidus, strain LGP32, is an oyster pathogen associated with the summer mortalities affecting the production of Crassostrea gigas oysters worldwide. Vibrio splendidus LGP32 was shown to resist to up to 10 µM Cg‐Def defensin and Cg‐BPI bactericidal permeability increasing protein, two antimicrobial peptides/proteins (AMPs) involved in C. gigas immunity. The resistance to both oyster Cg‐Def and Cg‐BPI and standard AMPs (polymyxin B, protegrin, human BPI) was dependent on the ompU gene. Indeed, upon ompU inactivation, minimal bactericidal concentrations decreased by up to fourfold. AMP resistance was restored upon ectopic expression of ompU. The susceptibility of bacterial membranes to AMP‐induced damages was independent of the ompU‐mediated AMP resistance. Besides its role in AMP resistance, ompU proved to be essential for the adherence of V. splendidus LGP32 to fibronectin. Interestingly, in vivo, ompU was identified as a major determinant of V. splendidus pathogenicity in oyster experimental infections. Indeed, the V. splendidus‐induced oyster mortalities dropped from 56% to 11% upon ompU mutation (Kaplan–Meier survival curves, P < 0.01). Moreover, in co‐infection assays, the ompU mutant was out competed by the wild‐type strain with competitive indexes in the range of 0.1–0.2. From this study, ompU is required for virulence of V. splendidus. Contributing to AMP resistance, conferring adhesive properties to V. splendidus, and being essential for in vivo fitness, the OmpU porin appears as an essential effector of the C. gigas/V. splendidus interaction.     

A Highlights from MBoC Selection: A Gαi–GIV Molecular Complex Binds Epidermal Growth Factor Receptor and Determines Whether Cells Migrate or Proliferate

Cells respond to growth factors by either migrating or proliferating, but not both at the same time, a phenomenon termed migration-proliferation dichotomy. The underlying mechanism of this phenomenon has remained unknown. We demonstrate here that Gα_i protein and GIV, its nonreceptor guanine nucleotide exchange factor (GEF), program EGF receptor (EGFR) signaling and orchestrate this dichotomy. GIV directly interacts with EGFR, and when its GEF function is intact, a Gα_i–GIV–EGFR signaling complex assembles, EGFR autophosphorylation is enhanced, and the receptor''s association with the plasma membrane (PM) is prolonged. Accordingly, PM-based motogenic signals (PI3-kinase-Akt and PLCγ1) are amplified, and cell migration is triggered. In cells expressing a GEF-deficient mutant, the Gαi–GIV-EGFR signaling complex is not assembled, EGFR autophosphorylation is reduced, the receptor''s association with endosomes is prolonged, mitogenic signals (ERK 1/2, Src, and STAT5) are amplified, and cell proliferation is triggered. In rapidly growing, poorly motile breast and colon cancer cells and in noninvasive colorectal carcinomas in situ in which EGFR signaling favors mitosis over motility, a GEF-deficient splice variant of GIV was identified. In slow growing, highly motile cancer cells and late invasive carcinomas, GIV is highly expressed and has an intact GEF motif. Thus, inclusion or exclusion of GIV''s GEF motif, which activates Gαi, modulates EGFR signaling, generates migration-proliferation dichotomy, and most likely influences cancer progression.     

Bioactive hydroxyphenylpyrrole-dicarboxylic acids from a new marine Halomonas sp.: production and structure elucidation

The new marine Halomonas sp. strain GWS-BW-H8hM (DSM 17996) was found to produce 3-(4′-hydroxyphenyl)-4-phenylpyrrole-2,5-dicarboxylic acid (HPPD-1) and 3,4-bis(4′-hydroxy- phenyl)pyrrole-2,5-dicarboxylic acid (HPPD-2). In initial cultivations using marine broth, only low contents of these compounds have been isolated. Improving the conditions and growing the strain on artificial seawater supplemented with tryptone 10 g l⁻¹, yeast extract 5 g l⁻¹, l-tyrosine 0.6 g l⁻¹, glycine 1 g l⁻¹, and glucose 6 g l^-1, the growth-associated HPPD-1 and HPPD-2 production of a 40-l batch cultivation reached the amounts of 47 mg l⁻¹ and 116 mg l⁻¹, respectively, after 65 h. Both compounds showed potent anti-tumor-promoting activities.     

Novel polysaccharide–protein-based amphipathic formulations

Previous results showed that the cell-surface esterase from Acinetobacter venetianus RAG-1 enhances the emulsification properties of the polymeric bioemulsifier emulsan and its deproteinated derivative apoemulsan (Bach H, Berdichevsky Y, Gutnick D (2003) An exocellular protein from the oil-degrading microbe Acinetobacter venetianus RAG-1 enhances the emulsifying activity of the polymeric bioemulsifier emulsan. Appl Environ Microbiol 69:2608–2615). Here we show that in the presence of the his-tagged recombinant esterase from RAG-1, 18 different polysaccharides from microbial, plant, insect and synthetic sources formed hexadecane-in-water emulsions. Emulsifying activities were distributed over a 13-fold range from over 4800 U/mg protein/mg polysaccharide in the case of apoemulsan to 370 U/mg protein/mg polysaccharide in the case of alginic acid. The stability of the emulsions ranged between 95 and 58%. Emulsions formed in the presence of seven of the polysaccharides exhibited stabilities of over 80%. The esterase from A. calcoaceticus BD4, which shows sequence homology to the RAG-1 esterase, was inactive in emulsification enhancement. The sequence of the RAG-1 esterase was shown to contain two conserved peptide sequences previously shown to be implicated in carbohydrate/polysaccharide binding. A hypothetical model illustrating a possible mode of interaction between the esterase, the apoemulsan and the oil droplet is presented. The complex is presumed to generate a series of “coated” oil droplets which are restricted in their ability to coalesce resulting in a relatively stable emulsion.     

The area composita of adhering junctions connecting heart muscle cells of vertebrates. I. Molecular definition in intercalated disks of cardiomyocytes by immunoelectron microscopy of desmosomal proteins

Among sarcomeric muscles the cardiac muscle cells are unique by, inter alia, a systemic and extended cell-cell contact structure, the intercalated disk (ID), comprising frequent and closely spaced arrays of plaque-coated cell-cell adhering junctions (AJs). As some of these junctions may look somewhat like desmosomes and others like fasciae adhaerentes, the dogma has emerged in the literature that IDs contain - like epithelial cells - both kinds of AJs formed by - for the most - mutually exclusive molecular ensembles. This, however, is not the case. In comprehensive immunoelectron microscopic studies of mammalian (human, bovine, rat, mouse) and non-mammalian (chicken, amphibia, fishes) heart muscle tissues, we have localized major constituents of the desmosomal plaques of polar epithelia, desmoplakin, plakophilin-2 and plakoglobin, as well as the desmosomal cadherins, desmoglein Dsg2 and desmocollin Dsc2, in both kinds of ID AJs, independent of the specific morphological appearance. The desmosomal molecules are not restricted to the desmosome-like-looking junctions but can also be detected in junctions appearing similar to the zonula or fascia adhaerens structures. These AJs of cardiac ID are therefore subsumed under the collective term area composita. We discuss our results with respect to the importance of ID junction molecules for the formation, maintenance and function of the heart, particularly in relation to recent findings that deletions of - or mutations in - genes encoding such proteins can cause severe, sometimes lethal damages.