Mutations in IMPG2, Encoding Interphotoreceptor Matrix Proteoglycan 2, Cause Autosomal-Recessive Retinitis Pigmentosa

Retinitis pigmentosa (RP) is a heterogeneous group of inherited retinal diseases caused by progressive degeneration of the photoreceptor cells. Using autozygosity mapping, we identified two families, each with three affected siblings sharing large overlapping homozygous regions that harbored the IMPG2 gene on chromosome 3. Sequence analysis of IMPG2 in the two index cases revealed homozygous mutations cosegregating with the disease in the respective families: three affected siblings of Iraqi Jewish ancestry displayed a nonsense mutation, and a Dutch family displayed a 1.8 kb genomic deletion that removes exon 9 and results in the absence of seven amino acids in a conserved SEA domain of the IMPG2 protein. Transient transfection of COS-1 cells showed that a construct expressing the wild-type SEA domain is properly targeted to the plasma membrane, whereas the mutant lacking the seven amino acids appears to be retained in the endoplasmic reticulum. Mutation analysis in ten additional index cases that were of Dutch, Israeli, Italian, and Pakistani origin and had homozygous regions encompassing IMPG2 revealed five additional mutations; four nonsense mutations and one missense mutation affecting a highly conserved phenylalanine residue. Most patients with IMPG2 mutations showed an early-onset form of RP with progressive visual-field loss and deterioration of visual acuity. The patient with the missense mutation, however, was diagnosed with maculopathy. The IMPG2 gene encodes the interphotoreceptor matrix proteoglycan IMPG2, which is a constituent of the interphotoreceptor matrix. Our data therefore show that mutations in a structural component of the interphotoreceptor matrix can cause arRP.     

Synthesis of Aβ[1‐42] and its derivatives with improved efficiency

It has been proved that the principal component of senile plaques is aggregates of β‐amyloid peptide (Aβ) in cases of one of the most common forms of age‐related neurodegenerative disorders, Alzheimer's disease (AD). Although the synthetic methods for the synthesis of Aβ peptides have been developed since their first syntheses, Aβ[1‐42] is still problematic to prepare. The highly hydrophobic composition of Aβ[1‐42] results in aggregation between resin‐bound peptide chains or intrachain aggregation which leads to a decrease in the rates of deprotection and repetitive incomplete coupling reactions during 9‐flurenylmethoxycarbonyl (Fmoc) synthesis. In order to avoid aggregation and/or disrupt internal aggregation during stepwise Fmoc solid phase synthesis and to improve the quality of crude products, several attempts have been made. Since highly pure Aβ peptides in large quantities are used in biological experiments, we wanted to develop a method for a rational synthesis of human Aβ[1‐42] with high purity and adequate yield. This paper reports a convenient methodology with a novel solvent system for the synthesis of Aβ[1‐42], its N‐terminally truncated derivatives Aβ[4‐42] and Aβ[5‐42], and Aβ[1‐42] labeled with 7‐amino‐4‐methyl‐3‐coumarinylacetic acid (AMCA) at the N‐terminus using Fmoc strategy. The use of 10% anisole in Dimethylformamide/Dichloromethane (DMF/DCM) can substantially improve the purity and yield of crude Aβ[1‐42] and has been shown to be an optimal coupling condition for the synthesis of Aβ[1‐42]. Anisole is a cheap and simple aid in the synthesis of ‘difficult sequences’ where other solvents are less successful in the prevention of aggregation during the synthesis. Copyright © 2006 European Peptide Society and John Wiley & Sons, Ltd.     

Proteomic analysis of membrane microdomains derived from both failing and non-failing human hearts

Eukaryotic cells plasma membranes are organized into microdomains of specialized function such as lipid rafts and caveolae, with a specific lipid composition highly enriched in cholesterol and glycosphingolipids. In addition to their role in regulating signal transduction, multiple functions have been proposed, such as anchorage of receptors, trafficking of cholesterol, and regulation of permeability. However, an extensive understanding of their protein composition in human heart, both in failing and non-failing conditions, is not yet available. Membrane microdomains were isolated from left ventricular tissue of both failing (n = 15) and non-failing (n = 15) human hearts. Protein composition and differential protein expression was explored by comparing series of 2-D maps and subsequent identification by LC-MS/MS analysis. Data indicated that heart membrane microdomains are enriched in chaperones, cytoskeletal-associated proteins, enzymes and protein involved in signal transduction pathway. In addition, differential protein expression profile revealed that 30 proteins were specifically up- or down-regulated in human heart failure membrane microdomains. This study resulted in the identification of human heart membrane microdomain protein composition, which was not previously available. Moreover, it allowed the identification of multiple proteins whose expression is altered in heart failure, thus opening new perspectives to determine which role they may play in this disease.     

Synthesis and biological evaluation of new conformationally biased integrin ligands based on a tetrahydroazoninone scaffold

The synthesis of new conformationally biased cyclic pentapeptides, incorporating the RGD sequence, and built around a tetrahydroazoninone scaffold, is reported. They exhibit interesting activity towards integrin alphaVbeta3 and a remarkable selectivity in comparison with integrin alphaVbeta5.     

Comparison of the Hematological Profile of Elite Road Cyclists during the 2010 and 2012 GiroBio Ten-Day Stage Races and Relationships with Final Ranking

Cycling stage races are strenuous endurance events during which exercise-induced variations in hematological parameters are consistently observed. However, specific literature on such changes is scarce and published data have been derived from small samples of athletes. The aims of this study were: (1) to determine the hematological response to middle-term strenuous endurance; and (2) to determine whether a relationship exists between the athlete-specific hematological profile and final placement in a cycling stage race. The study population was male professional cyclists (n = 253) competing in the 2010 (n = 144) and 2012 (n = 109) GiroBio 10-day stage races. Blood draws taken before the start of the race, at mid-race, and at end-race were performed in strict compliance with academic and anti-doping pre-analytical warnings. Blood chemistry included white blood cell, red blood cell, hemoglobin concentration, hematocrit, mean corpuscular volume (MCV), mean hemoglobin content (MCH), mean corpuscular hemoglobin content (MCHC), platelets, and reticulocyte relative and absolute counts. Compared to baseline values, erythrocyte, hemoglobin, hematocrit, MCHC, platelet and reticulocyte counts were all consistently lower at mid-race, but returned to normal by race-end, while leukocytes were increased in the final phase. MCV increased during both events. MCH increased in the first part to then return to baseline in the 2012 race. The calculated OFF-score consistently decreased in the first half of the race before increasing, but remained lower than the baseline value. The trends of variation in hematological parameters were substantially similar in both events. There was an inverse, albeit weak, relationship between placement and erythrocyte, platelet, hemoglobin, hematocrit and OFF-score values in the 2010, but not in the 2012 race. In conclusion, the data confirm that, in this large series of elite road cyclists, the strenuous effort a rider sustains during a stage race induces appreciable changes in the hematological profile.     

New cerebrosides from Euphorbia peplis L.: antimicrobial activity evaluation

The less polar fraction of the methanolic extract from the plant Euphorbia peplis L. exhibited interesting antifungal and antitubercular activity. A complex mixture of four glucocerebrosides was responsible for this activity. Two new cerebrosides were isolated for the first time from Euphorbiaceae, 4 was assigned as 1-O-(beta-D-glucopyranosyl)-(2S,3S,4E,8E)-2N-[(2'R)-2'-hydroxy-hexadecanoyl]-4 (E), 8 (E)-octadecadiene-1,3-diol and 3 as the 1-O-(beta-D-glucopyranosyl)-(2S,3S,4R,8Z)-2N-[(2'R)-2'-hydroxytetracosanoyl]-8 (Z)-octadecene-1,3,4-triol. The structures were determined on the basis of chemical and spectroscopic evidences. Mass spectrometry of dimethyl disulfide derivatives was useful for the determination of the double-bond positions in the long-chain bases.     

HOCTAR database: a unique resource for microRNA target prediction

An Antarctic strain (NJ-7) of Chlorella vulgaris possesses the same 18S rRNA sequence as that of a temperate strain (UTEX259), but shows significantly higher freezing tolerance than the latter. Suppression subtractive hybridization (SSH) was performed to identify genes of intensified expression in NJ-7 relative to UTEX259. Among the genes identified, Ccor1 and Ccor2, co-organized in the same gene cluster Ccor1-Ccor2-Ccor1-Ccor2, showed much higher expression levels in NJ-7 than in UTEX259 at both 20°C and 4°C. As detected by Northern blot and Western blot analyses, the two genes were cold-inducible in NJ-7 but almost not expressed in UTEX259. Their encoded products are predicted to share 55.7% identity to each other and possess physicochemical characteristics similar to that of late embryogenesis abundant (LEA) proteins in plants. The purified recombinant Ccor1 and Ccor2 showed high heat-stability and could act as cryoprotectants to lactate dehydrogenase in vitro. Based on their expression patterns and protein characteristics, we propose that Ccor1 and Ccor2 are two novel LEA proteins and are related to the greatly enhanced freezing tolerance in the Antarctic strain.     

Versatile synthesis of Boc protected hydrazinoacetic acid and its application to the chemoselective ligation of TASP molecules

This paper describes a convenient synthesis of protected hydrazine derivatives, i.e. 1,2-bis-Boc-hydrazinoacetic acid, and its application for hydrazone ligation techniques in convergent template assembled synthetic protein (TASP) synthesis.     

NOX5 variants are functionally active in endothelial cells

NADPH oxidases have been identified as sources of reactive oxygen species (ROS) in vascular cells. In addition to the initially described enzyme containing gp91phox (NOX2), several homologues to NOX2 have been identified. Whereas NOX1, NOX2, and NOX4 are expressed in endothelial cells, a functional role of NOX5 containing additional N-terminal calcium-binding domains of varying sequences has not been reported in these cells. NOX5 protein was found in the endoplasmic reticulum of human microvascular endothelial cells (HMEC-1) and in the vascular wall. HMEC-1 cells expressed NOX5beta and NOX5delta as well as a variant lacking calcium-binding domains (NOX5S). NOX5beta and NOX5S increased basal ROS levels. Ionomycin exclusively enhanced NOX5beta-mediated ROS production. Although p22phox, when overexpressed, interacted with both NOX5 proteins, it was not essential for NOX5-mediated ROS production. NOX5 proteins stimulated endothelial cell proliferation and the formation of capillary-like structures whereas depletion of NOX5 by siRNA prevented these responses to thrombin. These data show that endothelial cells express different NOX5 variants including NOX5S lacking calcium-binding domains. NOX5 proteins are functional, promoting endothelial ROS production, proliferation, and the formation of capillary-like structures and contribute to the endothelial response to thrombin. These findings suggest that NOX5 variants play a novel role in controlling ROS-dependent processes in the vasculature.     

Characterization of the OFD1/Ofd1 genes on the human and mouse sex chromosomes and exclusion of Ofd1 for the Xpl mouse mutant

Oral-facial-digital type 1 (OFD1) syndrome is an X-linked dominant condition characterized by malformations of the face, oral cavity, and digits. The responsible gene, OFD1, maps to human Xp22 and has an unknown function. We isolated and characterized the mouse Ofd1 gene and showed that it is subject to X-inactivation, in contrast to the human gene. Furthermore, we excluded a role for Ofd1 in the pathogenesis of the spontaneous mouse mutant Xpl, which had been proposed as a mouse model for this condition. Comparative sequence analysis demonstrated that OFD1 is conserved among vertebrates and absent in invertebrates. This analysis allowed the identification of evolutionarily conserved domains in the protein. Finally, we report the identification of 18 apparently nonfunctional OFD1 copies, organized in repeat units on the human Y chromosome. These degenerate OFD1-Y genes probably derived from the ancestral Y homologue of the X-linked gene. The high level of sequence identity among the different units suggests that duplication events have recently occurred during evolution.