Ligands binding to the prion protein induce its proteolytic release with therapeutic potential in neurodegenerative proteinopathies.

The prion protein (PrPC) is a central player in neurodegenerative diseases, such as prion diseases or Alzheimer's disease. In contrast to disease-promoting cell surface PrPC, extracellular fragments act neuroprotective by blocking neurotoxic disease-associated protein conformers. Fittingly, PrPC release by the metalloprotease ADAM10 represents a protective mechanism. We used biochemical, cell biological, morphological, and structural methods to investigate mechanisms stimulating this proteolytic shedding. Shed PrP negatively correlates with prion conversion and is markedly redistributed in murine brain in the presence of prion deposits or amyloid plaques, indicating a sequestrating activity. PrP-directed ligands cause structural changes in PrPC and increased shedding in cells and organotypic brain slice cultures. As an exception, some PrP-directed antibodies targeting repetitive epitopes do not cause shedding but surface clustering, endocytosis, and degradation of PrPC. Both mechanisms may contribute to beneficial actions described for PrP-directed ligands and pave the way for new therapeutic strategies against currently incurable neurodegenerative diseases.

Inhibition of group-I metabotropic glutamate receptors protects against prion toxicity.

Prion infections cause inexorable, progressive neurological dysfunction and neurodegeneration. Expression of the cellular prion protein PrPC is required for toxicity, suggesting the existence of deleterious PrPC-dependent signaling cascades. Because group-I metabotropic glutamate receptors (mGluR1 and mGluR5) can form complexes with the cellular prion protein (PrPC), we investigated the impact of mGluR1 and mGluR5 inhibition on prion toxicity ex vivo and in vivo. We found that pharmacological inhibition of mGluR1 and mGluR5 antagonized dose-dependently the neurotoxicity triggered by prion infection and by prion-mimetic anti-PrPC antibodies in organotypic brain slices. Prion-mimetic antibodies increased mGluR5 clustering around dendritic spines, mimicking the toxicity of Aß oligomers. Oral treatment with the mGluR5 inhibitor, MPEP, delayed the onset of motor deficits and moderately prolonged survival of prion-infected mice. Although group-I mGluR inhibition was not curative, these results suggest that it may alleviate the neurological dysfunctions induced by prion diseases.

Dynamic micro-organization of P2X7 receptors revealed by PALM based single particle tracking.

Adenosine triphosphate (ATP)-gated P2X7 receptors (P2X7Rs) are members of the purinergic receptor family that are expressed in several cell types including neurons. A high concentration of ATP is required for the channel opening of P2X7Rs compared to other members of this receptor family. Recent work suggests that ATP binding to members of the P2X receptor family determines the diffusion and localization of these receptors on the plasma membrane of neurons. Here, we employed single particle tracking photoactivated localization microscopy (sptPALM) to study the diffusion and ATP-dependence of rat P2X7Rs. Dendra2-tagged P2X7Rs were transfected in hippocampal neurons and imaged on proximal dendrites. Our results suggest the presence of two populations of P2X7Rs within the extra-synaptic membrane: a population composed of rapidly diffusing receptors and one stabilized within nanoclusters (~100 nm diameter). P2X7R trajectories were rarely observed at synaptic sites. P2X7R mutations in the ATP-binding site (K64A) or the conserved phosphorylation site (K17A) resulted in faster- and slower-diffusing receptors, respectively. Furthermore, ATP differentially accelerated wild type and K17A-mutant receptors but not K64A-mutant receptors. Our results indicate that receptor conformation plays a critical role in regulating ATP-mediated changes in P2X7R diffusion and micro-organization.

Molecular Biomarkers: their significance and application in marine pollution monitoring.

This paper presents an overview of the significance of the use of molecular biomarkers as diagnostic and prognostic tools for marine pollution monitoring. In order to assess the impact of highly persistent pollutants such as polychlorinated biphenyls (PCB), polychlorinated dibenzo-dioxins (PCDD), polychlorinated dibenzo-furans (PCDF), polynuclear aromatic hydrocarbons (PAH), tributyltin (TBT) and other toxic metals on the marine ecosystem a suite of biomarkers are being extensively used worldwide. Among the various types of biomarkers, the following have received special attention: cytochrome P4501A induction, DNA integrity, acetylcholinesterase activity and metallothionein induction. These biomarkers are being used to evaluate exposure of various species of sentinel marine organisms (e.g. mussels, clams, oysters, snails, fishes, etc.) to and the effect of various contaminants (organic xenobiotics and metals) using different molecular approaches [biochemical assays, enzyme linked immuno-sorbent assays (ELISA), spectrophotometric, fluorometric measurement, differential pulsed polarography, liquid chromatography, atomic absorption spectrometry]. The induction of the biotransformation enzyme, cytochrome P4501A in fishes (Callionymus lyra, Limanda limanda, Serranus sp., Mullus barbatus) and mussels (Dreissena polymorpha) by various xenobiotic contaminants such as PCBs, PAHs, PCDs is used as a biomarker of exposure to such organic pollutants. The induction of cytochrome P4501A is involved in chemical carcinogenesis through catalysis of the covalent bonding of organic contaminants to a DNA strand leading to formation of DNA adduct. Measurement of the induction of cytochrome P4501A in terms of EROD (7-ethoxy resorufin O-deethylase) activity is successfully used as a potential biomarker of exposure to xenobiotic contaminants in marine pollution monitoring. In order to assess the impact of neurotoxic compounds on marine environment the evaluation of acetylcholinesterase activity in marine organisms is used as a biomarker of exposure to neurotoxic agents such as organophosphorus, carbamate pesticides etc. Metallothioneins (MTs) are induced by toxic metals such as Cd, Hg, and Cu by chelation through cysteine residues and are used in both vertebrates and invertebrates as a biomarker of metal exposure. The measurement of the levels of DNA integrity in marine organisms such as Sea stars (Asterias rubens) from the North Sea and the marine snails (Planaxis sulcatus) from the Arabian Sea along the Goa coast exposed to environmental xenobiotic contaminants clearly indicated the extent and the nature of pollution at the sampling sites along coastal environment.