Synaptic vesicle membrane fusion complex: action of clostridial neurotoxins on assembly.

Clostridial neurotoxins inhibit neurotransmitter release by selective and specific intracellular proteolysis of synaptobrevin/VAMP, synaptosomal-associated protein of 25 kDa (SNAP-25) or syntaxin. Here we show that in binary reactions synaptobrevin binds weakly to both SNAP-25 and syntaxin, and SNAP-25 binds to syntaxin. In the presence of all three components, a dramatic increase in the interaction strengths occurs and a stable sodium dodecyl sulfate-resistant complex forms. Mapping of the interacting sequences reveals that complex formation correlates with the presence of predicted alpha-helical structures, suggesting that membrane fusion involves intermolecular interactions via coiled-coil structures. Most toxins only attack the free, and not the complexed, proteins, and proteolysis of the proteins by different clostridial neurotoxins has distinct inhibitory effects on the formation of synaptobrevin-syntaxin-SNAP-25 complexes. Our data suggest that synaptobrevin, syntaxin and SNAP-25 associate into a unique stable complex that functions in synaptic vesicle exocytosis.     

Cooperative Recruitment of Dynamin and BIN/Amphiphysin/Rvs (BAR) Domain-containing Proteins Leads to GTP-dependent Membrane Scission

Dynamin mediates various membrane fission events, including the scission of clathrin-coated vesicles. Here, we provide direct evidence for cooperative membrane recruitment of dynamin with the BIN/amphiphysin/Rvs (BAR) proteins, endophilin and amphiphysin. Surprisingly, endophilin and amphiphysin recruitment to membranes was also dependent on binding to dynamin due to auto-inhibition of BAR-membrane interactions. Consistent with reciprocal recruitment in vitro, dynamin recruitment to the plasma membrane in cells was strongly reduced by concomitant depletion of endophilin and amphiphysin, and conversely, depletion of dynamin dramatically reduced the recruitment of endophilin. In addition, amphiphysin depletion was observed to severely inhibit clathrin-mediated endocytosis. Furthermore, GTP-dependent membrane scission by dynamin was dramatically elevated by BAR domain proteins. Thus, BAR domain proteins and dynamin act in synergy in membrane recruitment and GTP-dependent vesicle scission.     

The dynamin superfamily: universal membrane tubulation and fission molecules?

Dynamins are large GTPases that belong to a protein superfamily that, in eukaryotic cells, includes classical dynamins, dynamin-like proteins, OPA1, Mx proteins, mitofusins and guanylate-binding proteins/atlastins. They are involved in many processes including budding of transport vesicles, division of organelles, cytokinesis and pathogen resistance. With sequenced genomes from Homo sapiens, Drosophila melanogaster, Caenorhabditis elegans, yeast species and Arabidopsis thaliana, we now have a complete picture of the members of the dynamin superfamily from different organisms. Here, we review the superfamily of dynamins and their related proteins, and propose that a common mechanism leading to membrane tubulation and/or fission could encompass their many varied functions.     

Guided Self-Help Cognitive Behavioural Therapy for Depression in Primary Care: A Randomised Controlled Trial

Background

Access to Cognitive behavioural therapy (CBT) for depression is limited. One solution is CBT self-help books. Trial Objectives: To assess the impact of a guided self-help CBT book (GSH-CBT) on mood, compared to treatment as usual (TAU).Hypotheses:

1. GSH-CBT will have improved mood and knowledge of the causes and treatment of depression compared to the control receiving TAU
2. Guided self-help will be acceptable to patients and staff.

Methods and Findings

Participants: Adults attending seven general practices in Glasgow, UK with a BDI-II score of ≥14. 141 randomised to GSH-CBT and 140 to TAU. Interventions: RCT comparing ‘Overcoming Depression: A Five Areas Approach’ book plus 3–4 short face to face support appointments totalling up to 2 hours of guided support, compared with general practitioner TAU. Primary outcome: The BDI (II) score at 4 months. Numbers analysed: 281 at baseline, 203 at 4 months (primary outcome), 117 at 12 months. Outcome: Mean BDI-II scores were lower in the GSH-CBT group at 4 months by 5.3 points (2.6 to 7.9, p<0.001). At 4 and 12 months there were also significantly higher proportions of participants achieving a 50% reduction in BDI-II in the GSH-CBT arm. The mean support was 2 sessions with 42.7 minutes for session 1, 41.4 minutes for session 2 and 40.2 minutes of support for session 3. Adverse effects/Harms: Significantly less deterioration in mood in GSH-CBT (2.0% compared to 9.8% in the TAU group for BDI—II category change).

Limitations

Weaknesses: Our follow-up rate of 72.2% at 4 months is better than predicted but is poorer at 12 months (41.6%). In the GSH-CBT arm, around 50% of people attended 2 or fewer sessions. 22% failed to take up treatment.

Conclusions

GSH-CBT is substantially more effective than TAU.

Trial Registration

Controlled-Trials.com ISRCTN13475030     

Myostatin-deficient mice lose more skeletal muscle mass than wild-type controls during hindlimb suspension

Myostatin inhibits myogenesis. Therefore, we sought to determine if mice lacking the myostatin gene [Mstn(-/-)] would lose less muscle mass than wild-type mice during 7 days of hindlimb suspension (HS). Male Mstn(-/-) and wild-type (C57) mice were subjected to HS or served as ground-based controls (n = 6/group). Wild-type mice lost 8% of body mass and approximately 13% of wet mass from biceps femoris, quadriceps femoris, and soleus, whereas the mass of extensor digitorum longus (EDL) was unchanged after HS. Unexpectedly, Mstn(-/-) mice lost more body (13%, P < 0.05) and quadriceps femoris (17%, P < 0.05) mass than wild-type mice and lost 33% of EDL mass (P < 0.01) after HS. Protein expression of myostatin in biceps femoris and quadriceps femoris was not altered, whereas expression of MyoD, Myf-5, and myogenin increased in wild-type mice and tended to decrease in muscles of Mstn(-/-) mice. These data suggest that HS induced myogenesis in wild-type mice to counter atrophy, whereas myogenesis was not induced in Mstn(-/-) mice, thereby resulting in a greater loss of muscle mass.     

Temperature and precipitation at migratory grounds influence demographic trends of an Arctic‐breeding bird

Anthropogenic climate disruption, including temperature and precipitation regime shifts, has been linked to animal population declines since the mid‐20th century. However, some species, such as Arctic‐breeding geese, have thrived during this period. An increased understanding of how climate disruption might link to demographic rates in thriving species is an important perspective in quantifying the impact of anthropogenic climate disruption on the global state of nature. The Greenland barnacle goose (Branta leucopsis) population has increased tenfold in abundance since the mid‐20th century. A concurrent weather regime shift towards warmer, wetter conditions occurred throughout its range in Greenland (breeding), Ireland and Scotland (wintering) and Iceland (spring and autumn staging). The aim of this study was to determine the relationship between weather and demographic rates of Greenland barnacle geese to discern the role of climate shifts in the population trend. We quantified the relationship between temperature and precipitation and Greenland barnacle goose survival and productivity over a 50 year period from 1968 to 2018. We detected significant positive relationships between warmer, wetter conditions on the Icelandic spring staging grounds and survival. We also detected contrasting relationships between warmer, wetter conditions during autumn staging and survival and productivity, with warm, dry conditions being the most favourable for productivity. Survival increased in the latter part of the study period, supporting the possibility that spring weather regime shifts contributed to the increasing population trend. This may be related to improved forage resources, as warming air temperatures have been shown to improve survival rates in several other Arctic and northern terrestrial herbivorous species through indirect bottom‐up effects on forage availability.     

Evolving nature of the AP2 alpha-appendage hub during clathrin-coated vesicle endocytosis

Clathrin-mediated endocytosis involves the assembly of a network of proteins that select cargo, modify membrane shape and drive invagination, vesicle scission and uncoating. This network is initially assembled around adaptor protein (AP) appendage domains, which are protein interaction hubs. Using crystallography, we show that FxDxF and WVxF peptide motifs from synaptojanin bind to distinct subdomains on alpha-appendages, called 'top' and 'side' sites. Appendages use both these sites to interact with their binding partners in vitro and in vivo. Occupation of both sites simultaneously results in high-affinity reversible interactions with lone appendages (e.g. eps15 and epsin1). Proteins with multiple copies of only one type of motif bind multiple appendages and so will aid adaptor clustering. These clustered alpha(appendage)-hubs have altered properties where they can sample many different binding partners, which in turn can interact with each other and indirectly with clathrin. In the final coated vesicle, most appendage binding partners are absent and thus the functional status of the appendage domain as an interaction hub is temporal and transitory giving directionality to vesicle assembly.     

The Retinoblastoma Protein Is Linked to the Activation of Ras

The inner membrane-bound protein Ras integrates various extracellular signals that are subsequently communicated from the cytoplasm to the nucleus via the Raf/MEK/MAPK cascade. Here we show that the retinoblastoma protein pRb, previously reported to be a nuclear target of this pathway, can in turn influence the activation state of Ras. Rb-deficient fibroblasts display elevated levels (up to 30-fold) of activated Ras during G(1). Expression of wild-type pRb or a number of pRb mutants defective in E2F regulation reverses this effect. We provide evidence that the mid-G(1) activation of Ras in Rb-deficient cells, which occurs at the level of guanine nucleotide binding, differs from that of epidermal growth factor-induced stimulation of Ras, being dependent on protein synthesis. The aberrant levels of Ras activity associated with loss of pRb may be responsible for the differentiation defects in Rb-deficient cells, because suppression of Ras activity in Rb(-/-) fibroblasts restores the transactivation function of MyoD and the expression of a late marker of skeletal muscle differentiation. These data suggest that nuclear-cytoplasmic communication between pRb and Ras is bidirectional.