Chemokine CXCL4 interactions with extracellular matrix proteoglycans mediate widespread immune cell recruitment independent of chemokine receptors

Leukocyte recruitment from the vasculature into tissues is a crucial component of the immune system but is also key to inflammatory disease. Chemokines are central to this process but have yet to be therapeutically targeted during inflammation due to a lack of mechanistic understanding. Specifically, CXCL4 (Platelet Factor 4, PF4) has no established receptor that explains its function. Here, we use biophysical, in vitro, and in vivo techniques to determine the mechanism underlying CXCL4-mediated leukocyte recruitment. We demonstrate that CXCL4 binds to glycosaminoglycan (GAG) sugars on proteoglycans within the endothelial extracellular matrix, resulting in increased adhesion of leukocytes to the vasculature, increased vascular permeability, and non-specific recruitment of a range of leukocytes. Furthermore, GAG sulfation confers selectivity onto chemokine localization. These findings present mechanistic insights into chemokine biology and provide future therapeutic targets

Hybrid Mass Spectrometry Approaches to Determine How L-Histidine Feedback Regulates the Enzyzme MtATP-Phosphoribosyltransferase

MtATP-phosphoribosyltransferase (MtATP-PRT) is an enzyme catalyzing the first step of the biosynthesis of L-histidine in Mycobacterium tuberculosis, and proposed to be regulated via an allosteric mechanism. Native mass spectrometry (MS) reveals MtATP-PRT to exist as a hexamer. Conformational changes induced by L-histidine binding and the influence of buffer pH are determined with ion mobility MS, hydrogen deuterium exchange (HDX) MS, and analytical ultracentrifugation. The experimental collision cross-section (DTCCSHe) decreases from 76.6 to 73.5 nm2 upon ligand binding at pH 6.8, which correlates to the decrease in CCS calculated from crystal structures. No such changes in conformation were found at pH 9.0. Further detail on the regions that exhibit conformational change on L-histidine binding is obtained with HDX-MS experiments. On incubation with L-histidine, rapid changes are observed within domain III, and around the active site at longer times, indicating an allosteric effect

Civil society in Central and Eastern Europe: The ambivalent legacy of accession

Civil society organisations in Central and Eastern Europe (CEE) have remained weak players compared to their counterparts in established democracies. Given the particular incentives that the EU offered for the empowerment of non-state actors during pre-accession, it has often been assumed that EU intervention improved this situation. We argue that, instead, the EU's impact was highly ambivalent. Although the EU aid and EU-induced policy reform levelled the way for established actors' involvement in multilevel politics, it reinforced some of the barriers to development that the civil society organisations face in CEE. In particular, EU measures have failed to address the lack of sustainable income, of formalised interactions with the state and of grassroot support. Drawing on the experiences of trade unions and environmental groups, we show that this ambivalent 'legacy of accession' is due to an unfortunate interrelation between various, often implicit mechanisms of the EU's enlargement regime on one hand, and particular problems inherited from state socialism and transition on the other. Acta Politica (2010) 45, 41-69. doi: 10.1057/ap.2009.1