Fusion pore regulation by cAMP/Epac2 controls cargo release during insulin exocytosis
Abstract
Regulated exocytosis establishes a narrow fusion pore as initial aqueous connection to the extracellular space, through which small transmitter molecules such as ATP can exit. Co-release of polypeptides and hormones like insulin requires further expansion of the pore. There is evidence that pore expansion is regulated and can fail in diabetes and neurodegenerative disease. Here we report that the cAMP-sensor Epac2 (Rap-GEF4) controls fusion pore behavior by acutely recruiting two pore-restricting proteins, amisyn and dynamin-1, to the exocytosis site in insulin-secreting beta-cells. cAMP elevation restricts and slows fusion pore expansion and peptide release, but not when Epac2 is inactivated pharmacologically or in Epac2-/- (Rapgef4-/-) mice. Consistently, overexpression of Epac2 impedes pore expansion. Widely used antidiabetic drugs (GLP-1 receptor agonists and sulfonylureas) activate this pathway and thereby paradoxically restrict hormone release. We conclude that Epac2/cAMP controls fusion pore expansion and thus the balance of hormone and transmitter release during insulin granule exocytosis.
Data availability
Source data file has been provided for Fig 7. All raw data are available on the Dryad Digital Repository (https://doi.org/10.5061/dryad.6b604g8).
-
Data from: Fusion pore regulation by cAMP/Epac2 controls cargo release during insulin exocytosisDryad Digital Repository, doi:10.5061/dryad.6b604g8.
Article and author information
Author details
Funding
Swedish Research Council (2014-02575)
- Anders Tengholm
- Sebastian Barg
Norwegian Research Council
- Marit Bakke
Helse-Bergen
- Marit Bakke
Olga Jönssons stipend
- Alenka Guček
P O Zetterlingsstiftelse
- Alenka Guček
Swedish Research Council (2017-00956)
- Anders Tengholm
- Sebastian Barg
Swedich Research Council (2018-02871)
- Anders Tengholm
- Sebastian Barg
European Foundation for the Study of Diabetes
- Anders Tengholm
- Sebastian Barg
Diabetes Wellness Network Sweden
- Anders Tengholm
- Sebastian Barg
Swedish Diabetes Society
- Anders Tengholm
- Sebastian Barg
Swedish Society for Medical Research
- Nikhil R Gandasi
Hjärnfonden
- Sebastian Barg
NovoNordisk
- Nikhil R Gandasi
- Anders Tengholm
- Sebastian Barg
Family Ernfors Foundation
- Alenka Guček
- Anders Tengholm
- Sebastian Barg
European Foundation for the Study of Diabetes
- Nikhil R Gandasi
- Anders Tengholm
- Sebastian Barg
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Axel T Brunger, Stanford University, United States
Ethics
Animal experimentation: This study was performed in strict accordance with European and Swedish legislation, fundamental ethical principles and approved by the Regional Ethics Board Uppsala (license number 31, 1-32).
Version history
- Received: September 5, 2018
- Accepted: April 28, 2019
- Accepted Manuscript published: May 17, 2019 (version 1)
- Accepted Manuscript updated: May 20, 2019 (version 2)
- Version of Record published: June 10, 2019 (version 3)
Copyright
© 2019, Guček 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
-
- 2,129
- views
-
- 385
- downloads
-
- 34
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
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)
Further reading
-
- Cell Biology
- Chromosomes and Gene Expression
Heat stress is a major threat to global crop production, and understanding its impact on plant fertility is crucial for developing climate-resilient crops. Despite the known negative effects of heat stress on plant reproduction, the underlying molecular mechanisms remain poorly understood. Here, we investigated the impact of elevated temperature on centromere structure and chromosome segregation during meiosis in Arabidopsis thaliana. Consistent with previous studies, heat stress leads to a decline in fertility and micronuclei formation in pollen mother cells. Our results reveal that elevated temperature causes a decrease in the amount of centromeric histone and the kinetochore protein BMF1 at meiotic centromeres with increasing temperature. Furthermore, we show that heat stress increases the duration of meiotic divisions and prolongs the activity of the spindle assembly checkpoint during meiosis I, indicating an impaired efficiency of the kinetochore attachments to spindle microtubules. Our analysis of mutants with reduced levels of centromeric histone suggests that weakened centromeres sensitize plants to elevated temperature, resulting in meiotic defects and reduced fertility even at moderate temperatures. These results indicate that the structure and functionality of meiotic centromeres in Arabidopsis are highly sensitive to heat stress, and suggest that centromeres and kinetochores may represent a critical bottleneck in plant adaptation to increasing temperatures.
-
- Cell Biology
High-altitude polycythemia (HAPC) affects individuals living at high altitudes, characterized by increased red blood cells (RBCs) production in response to hypoxic conditions. The exact mechanisms behind HAPC are not fully understood. We utilized a mouse model exposed to hypobaric hypoxia (HH), replicating the environmental conditions experienced at 6000 m above sea level, coupled with in vitro analysis of primary splenic macrophages under 1% O2 to investigate these mechanisms. Our findings indicate that HH significantly boosts erythropoiesis, leading to erythrocytosis and splenic changes, including initial contraction to splenomegaly over 14 days. A notable decrease in red pulp macrophages (RPMs) in the spleen, essential for RBCs processing, was observed, correlating with increased iron release and signs of ferroptosis. Prolonged exposure to hypoxia further exacerbated these effects, mirrored in human peripheral blood mononuclear cells. Single-cell sequencing showed a marked reduction in macrophage populations, affecting the spleen’s ability to clear RBCs and contributing to splenomegaly. Our findings suggest splenic ferroptosis contributes to decreased RPMs, affecting erythrophagocytosis and potentially fostering continuous RBCs production in HAPC. These insights could guide the development of targeted therapies for HAPC, emphasizing the importance of splenic macrophages in disease pathology.