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Membrane rafts and its GPI anchored complement inhibitors regulate insulin secretion

Nagaraj, Vini LU (2015) In Lund University, Faculty of Medicine Doctoral Dissertation Series 2015:85.
Abstract
Type 2 diabetic patients usually exhibit an abnormal lipid profile. One such lipid that is constantly elevated in T2D is cholesterol. At the cellular level, cholesterol aids in tight packing of sphingolipids in certain regions of the plasma membrane and these specialized regions are termed ‘membrane rafts’, which act as signalling hubs. In pancreatic beta cells, these structures have been found to anchor important exocytotic proteins like VAMP2, syntaxin1A, SNAP25, Ca2+ channels and K+ channels and there are increasing number of evidences for its role in insulin secretory process. This thesis has aimed at unravelling the mechanisms by which membrane rafts regulate insulin exocytosis.

Pancreatic islets from type 2 diabetic human... (More)
Type 2 diabetic patients usually exhibit an abnormal lipid profile. One such lipid that is constantly elevated in T2D is cholesterol. At the cellular level, cholesterol aids in tight packing of sphingolipids in certain regions of the plasma membrane and these specialized regions are termed ‘membrane rafts’, which act as signalling hubs. In pancreatic beta cells, these structures have been found to anchor important exocytotic proteins like VAMP2, syntaxin1A, SNAP25, Ca2+ channels and K+ channels and there are increasing number of evidences for its role in insulin secretory process. This thesis has aimed at unravelling the mechanisms by which membrane rafts regulate insulin exocytosis.

Pancreatic islets from type 2 diabetic human donors show markedly decreased membrane raft staining intensity when compared to islets from healthy donors. Oxidation of membrane raft cholesterol using cholesterol oxidase (CO) significantly reduced membrane rafts, thereby disrupting its native structure and thus mimicking the diabetic condition. We have here shown that membrane raft disruption causes increased secretion of insulin under non-stimulatory conditions, depicting the loss of regulated insulin secretion. The increased basal insulin levels are accounted for by increased voltage-gated Ca2+ channel activity, which is further translated into increased influx of Ca2+ under basal conditions.

Membrane rafts contain two important GPI anchored proteins, CD59 and CD55 which are crucial inhibitors of the complement system. Our results suggest that CD59 is the most highly expressed complement gene in human and rat pancreatic islets, the expression of which is downregulated in diabetic rodents. Intracellular CD59 is found colocalized with insulin, VAMP2 and syntaxin1A and this association is increased by high glucose incubations. Cells lacking CD59 exhibit decreased insulin exocytosis, which is caused by the lost CD59-mediated pairing of the SNARE proteins VAMP2 and syntaxin1A. CD59 thus plays an important role in insulin secretion. The extracellular CD59 is further observed to be important for maintaining the structural integrity of membrane rafts.

CD55 is highly expressed in pancreatic islets and beta cells. Interestingly, in chronic hyperglycemia (diabetes), the expression of CD55 is further increased and is positively correlated with HbA1c. In fact, this is probably directly related to the hyperglycemic condition as glucose incubations regulate CD55 expression in human islets. In contrast to CD59, CD55 neither interacts with, nor regulates the insulin secretion machinery. However, CD55 shares the important capacity to regulate membrane rafts, which are disintegrated by silencing of CD55.

Taken together, this thesis has shown that membrane rafts regulate insulin secretion by controlling the activity of voltage-gated Ca2+ channels. Furthermore, membrane raft marker protein and complement inhibitor CD59 is involved in the exocytosis process. This action is mediated by intracellular assistance in pairing of exocytosis-regulating SNARE proteins. (Less)
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author
supervisor
opponent
  • Ullrich, Susanne, Professor
organization
publishing date
type
Thesis
publication status
published
subject
keywords
Membrane rafts, basal insulin secretion, regulated insulin secretion, CD59, CD55, SNARE proteins, insulin, pancreatic beta cells.
in
Lund University, Faculty of Medicine Doctoral Dissertation Series
volume
2015:85
pages
71 pages
publisher
Islet patophysiology, Clinical Sciences, Malmö
defense location
Lecture theatre Medelhavet, Wallenberglaboratoriet, Malmö Skåne University Hospital, Malmö, Sweden.
defense date
2015-09-04 09:00
ISSN
1652-8220
ISBN
978-91-7619-164-4
language
English
LU publication?
yes
id
0c279ffe-561b-4964-8b9c-c692655db84b (old id 7760994)
date added to LUP
2015-08-13 09:35:34
date last changed
2016-09-19 08:44:47
@phdthesis{0c279ffe-561b-4964-8b9c-c692655db84b,
  abstract     = {Type 2 diabetic patients usually exhibit an abnormal lipid profile. One such lipid that is constantly elevated in T2D is cholesterol. At the cellular level, cholesterol aids in tight packing of sphingolipids in certain regions of the plasma membrane and these specialized regions are termed ‘membrane rafts’, which act as signalling hubs. In pancreatic beta cells, these structures have been found to anchor important exocytotic proteins like VAMP2, syntaxin1A, SNAP25, Ca2+ channels and K+ channels and there are increasing number of evidences for its role in insulin secretory process. This thesis has aimed at unravelling the mechanisms by which membrane rafts regulate insulin exocytosis.<br/><br>
Pancreatic islets from type 2 diabetic human donors show markedly decreased membrane raft staining intensity when compared to islets from healthy donors. Oxidation of membrane raft cholesterol using cholesterol oxidase (CO) significantly reduced membrane rafts, thereby disrupting its native structure and thus mimicking the diabetic condition. We have here shown that membrane raft disruption causes increased secretion of insulin under non-stimulatory conditions, depicting the loss of regulated insulin secretion. The increased basal insulin levels are accounted for by increased voltage-gated Ca2+ channel activity, which is further translated into increased influx of Ca2+ under basal conditions. <br/><br>
Membrane rafts contain two important GPI anchored proteins, CD59 and CD55 which are crucial inhibitors of the complement system. Our results suggest that CD59 is the most highly expressed complement gene in human and rat pancreatic islets, the expression of which is downregulated in diabetic rodents. Intracellular CD59 is found colocalized with insulin, VAMP2 and syntaxin1A and this association is increased by high glucose incubations. Cells lacking CD59 exhibit decreased insulin exocytosis, which is caused by the lost CD59-mediated pairing of the SNARE proteins VAMP2 and syntaxin1A. CD59 thus plays an important role in insulin secretion. The extracellular CD59 is further observed to be important for maintaining the structural integrity of membrane rafts. <br/><br>
CD55 is highly expressed in pancreatic islets and beta cells. Interestingly, in chronic hyperglycemia (diabetes), the expression of CD55 is further increased and is positively correlated with HbA1c. In fact, this is probably directly related to the hyperglycemic condition as glucose incubations regulate CD55 expression in human islets. In contrast to CD59, CD55 neither interacts with, nor regulates the insulin secretion machinery. However, CD55 shares the important capacity to regulate membrane rafts, which are disintegrated by silencing of CD55.<br/><br>
Taken together, this thesis has shown that membrane rafts regulate insulin secretion by controlling the activity of voltage-gated Ca2+ channels. Furthermore, membrane raft marker protein and complement inhibitor CD59 is involved in the exocytosis process. This action is mediated by intracellular assistance in pairing of exocytosis-regulating SNARE proteins.},
  author       = {Nagaraj, Vini},
  isbn         = {978-91-7619-164-4},
  issn         = {1652-8220},
  keyword      = {Membrane rafts,basal insulin secretion,regulated insulin secretion,CD59,CD55,SNARE proteins,insulin,pancreatic beta cells.},
  language     = {eng},
  pages        = {71},
  publisher    = {Islet patophysiology, Clinical Sciences, Malmö},
  school       = {Lund University},
  series       = {Lund University, Faculty of Medicine Doctoral Dissertation Series},
  title        = {Membrane rafts and its GPI anchored complement inhibitors regulate insulin secretion},
  volume       = {2015:85},
  year         = {2015},
}