The HSP70 Molecular Chaperone Is Not Beneficial in a Mouse Model of α-synucleinopathy

BACKGROUND: Aggregation and misfolded alpha-synuclein is thought to be central in the pathogenesis of Parkinson's disease (PD). Heat-shock proteins (HSPs) that are involved in refolding and degradation processes could lower the aggregate load of alpha-synuclein and thus be beneficial in alpha-synucleinopathies. METHODOLOGY/PRINCIPAL FINDINGS: We co-overexpressed human A53T point-mutated alpha-synuclein and human HSP70 in mice, both under the control of Thy1 regulatory sequences. Behavior read-outs showed no beneficial effect of HSP70 expression in mice. In contrast, motor coordination, grip strength and weight were even worse in the alpha-synucleinopathy model in the presence of HSP70 overexpression. Biochemical analyses revealed no differences in alpha-synuclein oligomers/aggregates, truncations and phosphorylation levels and alpha-synuclein localization was unchanged in immunostainings. CONCLUSION/SIGNIFICANCE: Overexpressing HSP70 in a mouse model of alpha-synucleinopathy did not lower the toxic load of alpha-synuclein species and had no beneficial effect on alpha-synuclein-related motor deficits

Efflux pump inhibitors (EPIs) as new antimicrobial agents against Pseudomonas aeruginosa

Pseudomonas aeruginosa is an opportunistic human pathogen and one of the leading causes of nosocomial infections worldwide. The difficulty in treatment of pseudomonas infections arises from being multidrug resistant (MDR) and exhibits resistance to most antimicrobial agents due to the expression of different mechanisms overcoming their effects. Of these resistance mechanisms, the active efflux pumps in Pseudomonas aeruginosa that belong to the resistance nodulation division (RND) plays a very important role in extruding the antibiotics outside the bacterial cells providing a protective means against their antibacterial activity. Beside its role against the antimicrobial agents, these pumps can extrude biocides, detergents, and other metabolic inhibitors. It is clear that efflux pumps can be targets for new antimicrobial agents. Peptidomimetic compounds such as phenylalanine arginyl β-naphthylamide (PAβN) have been introduced as efflux pump inhibitors (EPIs); their mechanism of action is through competitive inhibition with antibiotics on the efflux pump resulting in increased intracellular concentration of antibiotic, hence, restoring its antibacterial activity. The advantage of EPIs is the difficulty to develop bacterial resistance against them, but the disadvantage is their toxic property hindering their clinical application. The structure activity relationship of these compounds showed other derivatives from PAβN that are higher in their activity with higher solubility in biological fluids and decreased toxicity level. This raises further questions on how can we compact Pseudomonas infections. Of particular importance, the recent resurgence in the use of older antibiotics such as polymyxins and probably applying stricter control measures in order to prevent their spread in clinical sittings

NS2 Protein of Hepatitis C Virus Interacts with Structural and Non-Structural Proteins towards Virus Assembly

Growing experimental evidence indicates that, in addition to the physical virion components, the non-structural proteins of hepatitis C virus (HCV) are intimately involved in orchestrating morphogenesis. Since it is dispensable for HCV RNA replication, the non-structural viral protein NS2 is suggested to play a central role in HCV particle assembly. However, despite genetic evidences, we have almost no understanding about NS2 protein-protein interactions and their role in the production of infectious particles. Here, we used co-immunoprecipitation and/or fluorescence resonance energy transfer with fluorescence lifetime imaging microscopy analyses to study the interactions between NS2 and the viroporin p7 and the HCV glycoprotein E2. In addition, we used alanine scanning insertion mutagenesis as well as other mutations in the context of an infectious virus to investigate the functional role of NS2 in HCV assembly. Finally, the subcellular localization of NS2 and several mutants was analyzed by confocal microscopy. Our data demonstrate molecular interactions between NS2 and p7 and E2. Furthermore, we show that, in the context of an infectious virus, NS2 accumulates over time in endoplasmic reticulum-derived dotted structures and colocalizes with both the envelope glycoproteins and components of the replication complex in close proximity to the HCV core protein and lipid droplets, a location that has been shown to be essential for virus assembly. We show that NS2 transmembrane region is crucial for both E2 interaction and subcellular localization. Moreover, specific mutations in core, envelope proteins, p7 and NS5A reported to abolish viral assembly changed the subcellular localization of NS2 protein. Together, these observations indicate that NS2 protein attracts the envelope proteins at the assembly site and it crosstalks with non-structural proteins for virus assembly

The Wnt-dependent signaling pathways as target in oncology drug discovery

Our current understanding of the Wnt-dependent signaling pathways is mainly based on studies performed in a number of model organisms including, Xenopus, Drosophila melanogaster, Caenorhabditis elegans and mammals. These studies clearly indicate that the Wnt-dependent signaling pathways are conserved through evolution and control many events during embryonic development. Wnt pathways have been shown to regulate cell proliferation, morphology, motility as well as cell fate. The increasing interest of the scientific community, over the last decade, in the Wnt-dependent signaling pathways is supported by the documented importance of these pathways in a broad range of physiological conditions and disease states. For instance, it has been shown that inappropriate regulation and activation of these pathways is associated with several pathological disorders including cancer, retinopathy, tetra-amelia and bone and cartilage disease such as arthritis. In addition, several components of the Wnt-dependent signaling pathways appear to play important roles in diseases such as Alzheimer’s disease, schizophrenia, bipolar disorder and in the emerging field of stem cell research. In this review, we wish to present a focused overview of the function of the Wnt-dependent signaling pathways and their role in oncogenesis and cancer development. We also want to provide information on a selection of potential drug targets within these pathways for oncology drug discovery, and summarize current data on approaches, including the development of small-molecule inhibitors, that have shown relevant effects on the Wnt-dependent signaling pathways

Outer membrane protein folding from an energy landscape perspective

The cell envelope is essential for the survival of Gram-negative bacteria. This specialised membrane is densely packed with outer membrane proteins (OMPs), which perform a variety of functions. How OMPs fold into this crowded environment remains an open question. Here, we review current knowledge about OFMP folding mechanisms in vitro and discuss how the need to fold to a stable native state has shaped their folding energy landscapes. We also highlight the role of chaperones and the β-barrel assembly machinery (BAM) in assisting OMP folding in vivo and discuss proposed mechanisms by which this fascinating machinery may catalyse OMP folding