Cleavage of von Willebrand factor by granzyme M destroys its factor VIII binding capacity

Von Willebrand factor (VWF) is a pro-hemostatic multimeric plasma protein that promotes platelet aggregation and stabilizes coagulation factor VIII (FVIII) in plasma. The metalloproteinase ADAMTS13 regulates the platelet aggregation function of VWF via proteolysis. Severe deficiency of ADAMTS13 is associated with thrombotic thrombocytopenic purpura, but does not always correlate with its clinical course. Therefore, other proteases could also be important in regulating VWF activity. In the present study, we demonstrate that VWF is cleaved by the cytotoxic lymphocyte granule component granzyme M (GrM). GrM cleaved both denaturated and soluble plasma-derived VWF after Leu at position 276 in the D3 domain. GrM is unique in that it did not affect the multimeric size and pro-hemostatic platelet aggregation ability of VWF, but instead destroyed the binding of VWF to FVIII in vitro. In meningococcal sepsis patients, we found increased plasma GrM levels that positively correlated with an increased plasma VWF/FVIII ratio in vivo. We conclude that, next to its intracellular role in triggering apoptosis, GrM also exists extracellularly in plasma where it could play a physiological role in controlling blood coagulation by determining plasma FVIII levels via proteolytic processing of its carrier VWF.     

Trimeridianellidae nov. fam., a Triassic spumellarian radiolarian family with simple initial skeleton and pyloniacean mode of growth

Based on several samples of Middle and Late Triassic age, the authors describe a new family of spumellarian Radiolaria – Trimeridianellidae Dumitrica and Tekin, characterized by a three-fold symmetry, a first shell of triangular prism type with 6 primary rays originated at the edges of two bases of the prism, or 3 meridian arches with 3 equatorial rays, and growth of pyloniacean type. The family comprises 4 monospecific new genera (Enneacladus Dumitrica, Endoprisma Dumitrica, Trimeridianella Dumitrica and Tekin, and Tristylopyle Dumitrica), 4 new species and one left in open nomenclature. In spite of their spotty occurrences and low diversity, the authors attempt to trace the evolution of the family, which is considered to be the result of heterochrony in ontogenetic growth, mostly of paedomorphic type. The family seems to be related to the Middle Triassic family Patruliidae Dumitrica with which it shares in common the simple microsphere with 6 primary rays and the pyloniacean mode of growth.     

Neither lipophilicity nor membrane-perturbing potency of phenothiazine maleates correlate with the ability to inhibit P-glycoprotein transport activity

Although phenothiazines are known as multidrug resistance modifiers, the molecular mechanism of their activity remains unclear. Since phenothiazine molecules are amphiphilic, the interactions with membrane lipids may be related, at least partially, to their biological effects. Using the set of phenothiazine maleates differing in the type of phenothiazine ring substitution at position 2 and/or in the length of the alkyl bridge-connecting ring system and side chain group, we investigated if their ability to modulate the multidrug resistance of cancer cells correlated with model membrane perturbing potency. The influence exerted on lipid bilayers was determined by liposome/buffer partition coefficient measurements (using the absorption spectra second-derivative method), fluorescence spectroscopy and calorimetry. Biological effects were assessed by a flow cytometric functional test based on differential accumulation of fluorescent probe DiOC(2)(3) by parental and drug-resistant cells. We found that all phenothiazine maleates were incorporated into lipid bilayers and altered their biophysical properties. With only few exceptions, the extent of membrane perturbation induced by phenothiazine maleates correlated with their lipophilicity. Within the group of studied derivatives, the compounds substituted with CF(3)- at position 2 of phenothiazine ring were the most active membrane perturbants. No clear relation was found between effects exerted by phenothiazine maleates on model membranes and their ability to modulate P-glycoprotein transport activity.     

Citizenship, migration and formations of class in urban France

This paper is an exploration of the configurations of citizenship that prevail in Paris, a globalizing city where the processes of capitalist change and transnational migration converge. I will focus on the ways that different class segments of its migrant population exercise citizenship in a context in which the ideologies of belonging and membership are being redrawn under the demands of neoliberal transformation. My argument is that efforts made to rework models of citizenship under neoliberalism contribute not only to the realignments of class, but they also galvanise ethnic divisions and sentiments of nativism in France and more broadly in Europe. In making this argument, I draw on the notion of citizenship regime to focus attention on the political, economic and ideological forces that condition the orientations of the state, policies and citizenry in the context of crises and change under capitalism while also problematizing the state capital nexus in relation to the formation of subjects as citizens. This article is a continuation of a larger scholarly project that seeks to explore the ways in which the analytical paradigms of political economy advance our understanding of the different dimensions of migration and capitalist change.     

Characterization and optimization of β-galactosidase immobilization process on a mixed-matrix membrane

β-Galactosidase is an important enzyme catalyzing not only the hydrolysis of lactose to the monosaccharides glucose and galactose but also the transgalactosylation reaction to produce galacto-oligosaccharides (GOS). In this study, β-galactosidase was immobilized by adsorption on a mixed-matrix membrane containing zirconium dioxide. The maximum β-galactosidase adsorbed on these membranes was 1.6 g/m2, however, maximal activity was achieved at an enzyme concentration of around 0.5 g/m2. The tests conducted to investigate the optimal immobilization parameters suggested that higher immobilization can be achieved under extreme parameters (pH and temperature) but the activity was not retained at such extreme operational parameters. The investigations on immobilized enzymes indicated that no real shift occurred in its optimal temperature after immobilization though the activity in case of immobilized enzyme was better retained at lower temperature (5 °C). A shift of 0.5 unit was observed in optimal pH after immobilization (pH 6.5 to 7). Perhaps the most striking results are the kinetic parameters of the immobilized enzyme; while the Michaelis constant (K(m)) value increased almost eight times compared to the free enzyme, the maximum enzyme velocity (V(max)) remained almost constant.     

The influence of cyclomaltooligosaccharides (cyclodextrins) on the enzymatic decomposition of l-phenylalanine catalyzed by phenylalanine ammonia-lyase

Cyclomaltohexaose (α-cyclodextrin) and cyclomaltoheptaose (β-cyclodextrin) as well as their four methyl ether derivatives, that is, hexakis(2,3-di-O-methyl)cyclomaltohexaose, hexakis(2,3,6-tri-O-methyl)cyclomaltohexaose, heptakis(2,3-di-O-methyl)cyclomaltoheptaose, and heptakis(2,3,6-tri-O-methyl)cyclomaltoheptaose were investigated as the additives in the course of enzymatic decomposition of l-phenylalanine catalyzed by phenylalanine ammonia-lyase. Only a few of those additives behaved like classical inhibitors of the enzymatic reaction under investigation because the values of the Michaelis constants that were obtained, as well as the maximum velocity values depended mostly atypically on the concentrations of those additives. In most cases cyclodextrins caused mixed inhibition, both competitive and noncompetitive, but they also acted as activators for selected concentrations. This atypical behaviour of cyclodextrins is caused by three different and independent effects. The inhibitory effect of cyclodextrins is connected with the decrease of substrate concentration and unfavourable influence on the flexibility of the enzyme molecules. On the other hand, the activating effect is connected with the decrease of product concentration (the product is an inhibitor of the enzymatic reaction under investigation). All these effects are caused by the ability of the cyclodextrins to form inclusion complexes.     

A unique region of RILP distinguishes it from its related proteins in its regulation of lysosomal morphology and interaction with Rab7 and Rab34

Rab7 and Rab34 are implicated in regulation of lysosomal morphology and they share a common effector referred to as the RILP (Rab-interacting lysosomal protein). Two novel proteins related to RILP were identified and are tentatively referred to as RLP1 and RLP2 (for RILP-like protein 1 and 2, respectively). Overexpression of RILP caused enlarged lysosomes that are positioned more centrally in the cell. However, the morphology and distribution of lysosomes were not affected by overexpression of either RLP1 or RLP2. The molecular basis for the effect of RILP on lysosomes was investigated, leading to the demonstration that a 62-residue region (amino acids 272-333) of RILP is necessary for RILP's role in regulating lysosomal morphology. Remarkably, transferring this 62-residue region unique to RILP into corresponding sites in RLP1 rendered the chimeric protein capable of regulating lysosome morphology. A correlation between the interaction with GTP-bound form of both Rab proteins and the capability of regulating lysosomes was established. These results define a unique region in RILP responsible for its specific role in regulating lysosomal morphology as well as in its interaction with Rab7 and Rab34.