Project Details
Projekt
Dissecting the role of the scaffold proteins intersectin 1 and intersectin 2 in insulin secretion
Subject Area
Cell Biology
Term
from 2018 to 2023
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 407395267
The secretion of insulin by pancreatic b-cells is essential for glucose homeostasis. Defects in insulin release are a major factor in the pathogenesis of type 2 diabetes (T2D), a rising health problem. Insulin is stored in large granules to be released upon increased glucose concentration. Insulin secretion requires a well-orchestrated sequence of events comprising granule transport to the plasma membrane, remodeling of the local actin cytoskeleton, granule docking and finally fusion which has to be followed by endocytosis, potentially to retrieve critical components for subsequent release events. While the number of proteins known to participate in insulin secretion is growing, important details of the listed processes and especially the mechanisms underlying their spatiotemporal coordination remain elusive. Within this proposal we will tackle the fundamental question of how the different steps of insulin secretion are orchestrated. We hypothesize that the intersectin proteins, ITSN1 and ITSN2, both abundantly expressed in b-cells, are prime candidates for this role, since they are perfectly equipped for scaffolding and regulating large macromolecular assemblies due to their numerous interaction domains, and since they are known to serve as an interface between vesicle exo-/endocytosis and actin dynamics in neurotransmitter release. This hypothesis is especially appealing since ITSN1 transcripts are actually lower in b-cells from diabetic mice and patients suggesting that altered ITSN functionality might contribute to the pathomechanism of T2D. In fact, our experiments using ITSN1/2 knockout (KO) mice reveal a strongly decreased insulin secretion upon loss of ITSNs in vitro and in vivo. In addition, they indicate that ITSN1/2 deficient b-cells display impaired endocytosis which likely contributes to the reduced insulin secretion. This provides us with an excellent tool to dissect the so far unclear mechanism by which impaired endocytosis compromises insulin release. Thus, the aim of our proposal is to analyze the physiological importance of ITSN1/2 for insulin secretion, and to clarify the mechanistic basis for the influence of ITSN1/2 on insulin release by characterizing beta-cell specific ITSN1/2 KO mice. More specifically, using advanced live cell imaging, high pressure freezing electron microscopy, biochemistry, mutagenesis and mass spectrometry we will dissect whether the ITSNs act at the level of regulation of actin dynamics, vesicle docking, exocytosis and/or endocytosis, and whether they are needed for the overall orchestration of these processes. Finally, we will address how the ITSNs are regulated in insulin secretion and which role their altered expression plays in the pathogenesis of T2D. Overall, our studies will provide important insights into the mechanics and spatiotemporal coordination of the different processes underlying insulin secretion, with a special focus on the elusive role and regulation of endocytosis in insulin release.
DFG Programme
Research Grants
