A proline-rich motif on VGLUT1 reduces synaptic vesicle super-pool and spontaneous release frequency
- CNRS - Université de Bordeaux, France
- CNRS - INSERM - Université Pierre et Marie Curie, France
- Institut Pasteur, France
- CNRS, Université Paris-Sud, Université Paris-Saclay, France
- Max-Planck-Institut fuer Experimentelle Medizin, Germany
Abstract
Glutamate secretion at excitatory synapses is tightly regulated to allow for the precise tuning of synaptic strength. Vesicular Glutamate Transporters (VGLUT) accumulate glutamate into synaptic vesicles (SV) and thereby regulate quantal size. Further, the number of release sites and the release probability of SVs maybe regulated by the organization of active zone proteins and SV clusters. In the present work, we uncover a mechanism mediating an increased SV clustering through the interaction of VGLUT1 second proline-rich domain, endophilinA1 and intersectin1. This strengthening of SV clusters results in a combined reduction of axonal SV super-pool size and miniature excitatory events frequency. Our findings support a model in which clustered vesicles are held together through multiple weak interactions between Src homology 3 and proline-rich domains of synaptic proteins. In mammals, VGLUT1 gained a proline-rich sequence that recruits endophilinA1 and turns the transporter into a regulator of SV organization and spontaneous release.
Data availability
Raw measures and intermediate data processing of images and electrophysiology traces are submited in source files appended to this submission.Source images and electrophysiology traces reported in this study are fully available upon request to the corresponding author (Etienne Herzog - https://orcid.org/0000-0002-0058-6959).
Article and author information
Author details
Fabrice P Cordelières
Funding
Agence Nationale de la Recherche (ANR-12-JSV4-0005-01 VGLUT-IQ)
- Etienne Herzog
Agence Nationale de la Recherche (ANR-10-LABX-43 BRAIN)
- Yann Humeau
- Etienne Herzog
Agence Nationale de la Recherche (ANR-10-IDEX-03-02 PEPS SV-PIT)
- Etienne Herzog
Agence Nationale de la Recherche (PSYVGLUT 09-MNPS-033)
- Salah El Mestikawy
Agence Nationale de la Recherche (ANR-10-INBS-04)
- Fabrice P Cordelières
European Commission (Erasmus-Mundus European Neuroscience Campus Program)
- Xiao Min Zhang
- Etienne Herzog
Fondation pour la Recherche Médicale (FDT20120925288)
- Kätlin Silm
Fondation pour la Recherche Médicale (ING20150532192)
- Maria Florencia Angelo
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: The experimental design and all procedures were in accordance with the European guide for the care and use of laboratory animals, approved by the ethics committee of Bordeaux Universities (CE50), and registered with the french ministry for research under the APAFIS n{degree sign}1692. Every effort was made to minimize the number of animals used and their suffering.
Copyright
© 2019, Zhang et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 2,474
- views
-
- 397
- downloads
-
- 17
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
A proline-rich motif on VGLUT1 reduces synaptic vesicle super-pool and spontaneous release frequency
eLife 8:e50401.
https://doi.org/10.7554/eLife.50401
Further reading
-
- Cancer Biology
- Cell Biology
Understanding the cell cycle at the single-cell level is crucial for cellular biology and cancer research. While current methods using fluorescent markers have improved the study of adherent cells, non-adherent cells remain challenging. In this study, we addressed this gap by combining a specialized surface to enhance cell attachment, the FUCCI(CA)2 sensor, an automated image analysis pipeline, and a custom machine learning algorithm. This approach enabled precise measurement of cell cycle phase durations in non-adherent cells. This method was validated in acute myeloid leukemia cell lines NB4 and Kasumi-1, which have unique cell cycle characteristics, and we tested the impact of cell cycle-modulating drugs on NB4 cells. Our cell cycle analysis system, which is also compatible with adherent cells, is fully automated and freely available, providing detailed insights from hundreds of cells under various conditions. This report presents a valuable tool for advancing cancer research and drug development by enabling comprehensive, automated cell cycle analysis in both adherent and non-adherent cells.
-
- Cell Biology
Multiple gut antimicrobial mechanisms are coordinated in space and time to efficiently fight foodborne pathogens. In Drosophila melanogaster, production of reactive oxygen species (ROS) and antimicrobial peptides (AMPs) together with intestinal cell renewal play a key role in eliminating gut microbes. A complementary mechanism would be to isolate and treat pathogenic bacteria while allowing colonization by commensals. Using real-time imaging to follow the fate of ingested bacteria, we demonstrate that while commensal Lactiplantibacillus plantarum freely circulate within the intestinal lumen, pathogenic strains such as Erwinia carotovora or Bacillus thuringiensis, are blocked in the anterior midgut where they are rapidly eliminated by antimicrobial peptides. This sequestration of pathogenic bacteria in the anterior midgut requires the Duox enzyme in enterocytes, and both TrpA1 and Dh31 in enteroendocrine cells. Supplementing larval food with hCGRP, the human homolog of Dh31, is sufficient to block the bacteria, suggesting the existence of a conserved mechanism. While the immune deficiency (IMD) pathway is essential for eliminating the trapped bacteria, it is dispensable for the blockage. Genetic manipulations impairing bacterial compartmentalization result in abnormal colonization of posterior midgut regions by pathogenic bacteria. Despite a functional IMD pathway, this ectopic colonization leads to bacterial proliferation and larval death, demonstrating the critical role of bacteria anterior sequestration in larval defense. Our study reveals a temporal orchestration during which pathogenic bacteria, but not innocuous, are confined in the anterior part of the midgut in which they are eliminated in an IMD-pathway-dependent manner.
