Munc18-1: sequential interactions with the fusion machinery stimulate vesicle docking and priming

Exocytosis of secretory or synaptic vesicles is executed by a mechanism including the SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins. Munc18-1 is a part of this fusion machinery, but its role is controversial because it is indispensable for fusion but also inhibits the assembly of purified SNAREs in vitro. This inhibition reflects the binding of Munc18-1 to a closed conformation of the target-SNARE syntaxin1. The controversy would be solved if binding to closed syntaxin1 were shown to be stimulatory for vesicle fusion and/or additional essential interactions were identified between Munc18-1 and the fusion machinery. Here, we provide evidence for both notions by dissecting sequential steps of the exocytotic cascade while expressing Munc18 variants in the Munc18-1 null background. In Munc18-1 null chromaffin cells, vesicle docking is abolished and syntaxin levels are reduced. A mutation that diminished Munc18 binding to syntaxin1 in vitro attenuated the vesicle-docking step but rescued vesicle priming in excess of docking. Conversely, expressing the Munc18-2 isoform, which also displays binding to closed syntaxin1, rescued vesicle docking identical with Munc18-1 but impaired more downstream vesicle priming steps. All Munc18 variants restored syntaxin1 levels at least to wild-type levels, showing that the docking phenotype is not caused by syntaxin1 reduction. None of the Munc18 variants affected vesicle fusion kinetics or fusion pore duration. In conclusion, binding of Munc18-1 to closed syntaxin1 stimulates vesicle docking and a distinct interaction mode regulates the consecutive priming step. Copyright © 2007 Society for Neuroscience

A Munc18-1 mutant mimicking phosphorylation by Down Syndrome-related kinase Dyrk1a supports normal synaptic transmission and promotes recovery after intense activity

Phosphorylation of Munc18-1 (Stxbp1), a presynaptic organizer of synaptic vesicle fusion, is a powerful mechanism to regulate synaptic strength. Munc18-1 is a proposed substrate for the Down Syndrome-related kinase dual-specificity tyrosine phosphorylation-regulate kinase 1a (Dyrk1a) and mutations in both genes cause intellectual disability. However, the functional consequences of Dyrk1a-dependent phosphorylation of Munc18-1 for synapse function are unknown. Here, we show that the proposed Munc18-1 phosphorylation site, T479, is among the highly constrained phosphorylation sites in the coding regions of the gene and is also located within a larger constrained coding region. We confirm that Dyrk1a phosphorylates Munc18-1 at T479. Patch-clamp physiology in conditional null mutant hippocampal neurons expressing Cre and either wildtype, or mutants mimicking or preventing phosphorylation, revealed that synaptic transmission is similar among the three groups: frequency/amplitude of mEPSCs, evoked EPSCs, paired pulse plasticity, rundown kinetics upon intense activity and the readily releasable pool. However, synapses expressing the phosphomimic mutant responded to intense activity with more pronounced facilitation. These data indicate that Dyrk1a-dependent Munc18-1 phosphorylation has a minor impact on synaptic transmission, only after intense activity, and that the role of genetic variation in both genes in intellectual disability may be through different mechanisms

The RAB3-RIM Pathway Is Essential for the Release of Neuromodulators

Neurons secrete neuromodulators/neuropeptides from dense-core vesicles (DCVs) by a largely unknown mechanism. Persoon et al. identify RAB3 and RIM1/2 as essential factors. RAB3’s indispensable role is the first distinct feature of DCV secretion as compared to synaptic vesicle secretion

The Collaboration on Attachment Transmission Synthesis (CATS): A Move to the Level of Individual-Participant-Data Meta-Analysis.

Generations of researchers have tested and used attachment theory to understand children's development. To bring coherence to the expansive set of findings from small-sample studies, the field early on adopted meta-analysis. Nevertheless, gaps in understanding intergenerational transmission of individual differences in attachment continue to exist. We discuss how attachment research has been addressing these challenges by collaborating in formulating questions and pooling data and resources for individual-participant-data meta-analyses. The collaborative model means that sharing hard-won and valuable data goes hand in hand with directly and intensively interacting with a large community of researchers in the initiation phase of research, deliberating on and critically reviewing new hypotheses, and providing access to a large, carefully curated pool of data for testing these hypotheses. Challenges in pooling data are also discussed.Wellcom