A role for an Hsp70 nucleotide exchange factor in the regulation of synaptic vesicle endocytosis

© The Author(s), 2013. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Journal of Neuroscience 33 (2013): 8009-8021, doi:10.1523/JNEUROSCI.4505-12.2013.Neurotransmission requires a continuously available pool of synaptic vesicles (SVs) that can fuse with the plasma membrane and release their neurotransmitter contents upon stimulation. After fusion, SV membranes and membrane proteins are retrieved from the presynaptic plasma membrane by clathrin-mediated endocytosis. After the internalization of a clathrin-coated vesicle, the vesicle must uncoat to replenish the pool of SVs. Clathrin-coated vesicle uncoating requires ATP and is mediated by the ubiquitous molecular chaperone Hsc70. In vitro, depolymerized clathrin forms a stable complex with Hsc70*ADP. This complex can be dissociated by nucleotide exchange factors (NEFs) that release ADP from Hsc70, allowing ATP to bind and induce disruption of the clathrin:Hsc70 association. Whether NEFs generally play similar roles in vesicle trafficking in vivo and whether they play such roles in SV endocytosis in particular is unknown. To address this question, we used information from recent structural and mechanistic studies of Hsp70:NEF and Hsp70:co-chaperone interactions to design a NEF inhibitor. Using acute perturbations at giant reticulospinal synapses of the sea lamprey (Petromyzon marinus), we found that this NEF inhibitor inhibited SV endocytosis. When this inhibitor was mutated so that it could no longer bind and inhibit Hsp110 (a NEF that we find to be highly abundant in brain cytosol), its ability to inhibit SV endocytosis was eliminated. These observations indicate that the action of a NEF, most likely Hsp110, is normally required during SV trafficking to release clathrin from Hsc70 and make it available for additional rounds of endocytosis.This work was supported by the National Institutes of Health (Grant #NS029051 to E.M.L. and Grant #NS078165 to J.R.M.).2013-11-0

Understanding how adherence goals promote adherence behaviours: a repeated measure observational study with HIV seropositive patients

Background The extent to which patients follow treatments as prescribed is pivotal to treatment success. An exceptionally high level (> 95 %) of HIV medication adherence is required to suppress viral replication and protect the immune system and a similarly high level (> 80 %) of adherence has also been suggested in order to benefit from prescribed exercise programmes. However, in clinical practice, adherence to both often falls below the desirable level. This project aims to investigate a wide range of psychological and personality factors that may lead to adherence/non-adherence to medical treatment and exercise programmes. Methods HIV positive patients who are referred to the physiotherapist-led 10-week exercise programme as part of the standard care are continuously recruited. Data on social cognitive variables (attitude, intention, subjective norms, self-efficacy, and outcome beliefs) about the goal and specific behaviours, selected personality factors, perceived quality of life, physical activity, self-reported adherence and physical assessment are collected at baseline, at the end of the exercise programme and again 3 months later. The project incorporates objective measures of both exercise (attendance log and improvement in physical measures such as improved fitness level, weight loss, improved circumferential anthropometric measures) and medication adherence (verified by non-invasive hair analysis). Discussion The novelty of this project comes from two key aspects, complemented with objective information on exercise and medication adherence. The project assesses beliefs about both the underlying goal such as following prescribed treatment; and about the specific behaviours such as undertaking the exercise or taking the medication, using both implicit and explicit assessments of patients' beliefs and attitudes. We predict that i) the way people think about the underlying goal of their treatments explains medication and exercise behaviours over and above the effects of the behaviour-specific thinking and ii) the relationship between adherence to exercise and to medical treatment is stronger among those with more favourable views about the goal. Results from this study should identify the key contributing factors to inform subsequent adherence research and afford a more streamlined assessment matrix. The project also aims to inform patient care practices. UK Clinical Research Network registration number: UKCRN 7842