Article thumbnail

β2-Syntrophin Is a Cdk5 Substrate That Restrains the Motility of Insulin Secretory Granules

By Sandra Schubert, Klaus-Peter Knoch, Joke Ouwendijk, Shabaz Mohammed, Yury Bodrov, Melanie Jäger, Anke Altkrüger, Carolin Wegbrod, Marvin E. Adams, Yong Kim, Stanley C. Froehner, Ole N. Jensen, Yannis Kalaidzidis and Michele Solimena

Abstract

The molecular basis for the interaction of insulin granules with the cortical cytoskeleton of pancreatic β-cells remains unknown. We have proposed that binding of the granule protein ICA512 to the PDZ domain of β2-syntrophin anchors granules to actin filaments and that the phosphorylation/dephosphorylation of β2-syntrophin regulates this association. Here we tested this hypothesis by analyzing INS-1 cells expressing GFP-β2-syntrophin through the combined use of biochemical approaches, imaging studies by confocal and total internal reflection fluorescence microscopy as well as electron microscopy. Our results support the notion that β2-syntrophin restrains the mobility of cortical granules in insulinoma INS-1 cells, thereby reducing insulin secretion and increasing insulin stores in resting cells, while increasing insulin release upon stimulation. Using mass spectrometry, in vitro phosphorylation assays and β2-syntrophin phosphomutants we found that phosphorylation of β2-syntrophin on S75 near the PDZ domain decreases its binding to ICA512 and correlates with increased granule motility, while phosphorylation of S90 has opposite effects. We further show that Cdk5, which regulates insulin secretion, phosphorylates S75. These findings provide mechanistic insight into how stimulation displaces insulin granules from cortical actin, thus promoting their motility and exocytosis

Topics: Research Article
Publisher: Public Library of Science
OAI identifier: oai:pubmedcentral.nih.gov:2944849
Provided by: PubMed Central

To submit an update or takedown request for this paper, please submit an Update/Correction/Removal Request.

Suggested articles

Citations

  1. (2007b) Myosin va mediates docking of secretory granules at the plasma membrane.
  2. (2002). A monomeric red fluorescent protein.
  3. (1999). A PDZ-containing scaffold related to the dystrophin complex at the basolateral membrane of epithelial cells.
  4. (1987). A postsynaptic Mr 58,000 (58K) protein concentrated at acetylcholine receptorrich sites in Torpedo electroplaques and skeletal muscle.
  5. (2001). A real-time view of life within 100 nm of the plasma membrane.
  6. (1993). A role for the dystrophin-glycoprotein complex as a transmembrane linker between laminin and actin.
  7. (2000). Actindependent propulsion of endosomes and lysosomes by recruitment of N-WASP.
  8. (2003). An alpha-syntrophin-dependent pool of AQP4 in astroglial end-feet confers bidirectional water flow between blood and brain.
  9. (2006). Analysis of transient behavior in complex trajectories: application to secretory vesicle dynamics.
  10. (2000). Aquaporin-4 is absent at the sarcolemma and at perivascular astrocyte endfeet in a ´1-syntrophin knockout mice.
  11. (2000). Association of cortactin with dynamic actin in lamellipodia and on endosomal vesicles.
  12. (1994). Association of utrophin and multiple dystrophin short forms with the mammalian M(r) 58,000 dystrophin-associated protein (syntrophin).
  13. (2000). Biochemical characterisation of the actinbinding properties of utrophin.
  14. (2006). cAMP-dependent phosphorylation of PTB1 promotes the expression of insulin secretory granule proteins in beta cells.
  15. (2003). Cdk5 and the mystery of synaptic vesicle endocytosis.
  16. (2003). Cdk5 is essential for synaptic vesicle endocytosis.
  17. (2005). Cdk5-dependent regulation of glucose-stimulated insulin secretion.
  18. (2002). Cdk5/p35 regulates neurotransmitter release through phosphorylation and downregulation of P/Q-type voltage-dependent calcium channel activity.
  19. (1992). Cloning and expression of IDDM-specific human autoantigens.
  20. (2005). Common principles in clathrin-mediated sorting at the Golgi and the plasma membrane.
  21. (2003). Cophosphorylation of amphiphysin I and dynamin I by Cdk5 regulates clathrinmediated endocytosis of synaptic vesicles.
  22. (1991). Cortical filamentous actin disassembly and scinderin redistribution during chromaffin cell stimulation precede exocytosis, a phenomenon not exhibited by gelsolin.
  23. (2001). Cyclin-dependent protein kinase 5 (Cdk5) and the regulation of neurofilament metabolism.
  24. (2004). Cyclindependent kinase 5 associated with p39 promotes Munc18-1 phosphorylation and Ca(2+)-dependent exocytosis.
  25. (2001). Dephosphorylation of beta2-syntrophin and Ca2+/mu-calpain-mediated cleavage of ICA512 upon stimulation of insulin secretion.
  26. (2003). Developmental regulation of the proteolysis of the p35 cyclin-dependent kinase 5 activator by phosphorylation.
  27. (2005). Effect of oxygenated perfluorocarbons on isolated rat pancreatic islets in culture.
  28. (2000). Elzagallaai A
  29. (1999). Endocytic vesicles move at the tips of actin tails in cultured mast cells.
  30. (1992). Establishment of 2-mercaptoethanol-dependent differentiated insulin-secreting cell lines.
  31. (2005). Functional analysis of slac2-a/ melanophilin as a linker protein between Rab27A and myosin Va in melanosome transport.
  32. (2000). Gamma1- and gamma2-syntrophins, two novel dystrophin-binding proteins localized in neuronal cells.
  33. (2006). Global, in vivo, and site-specific phosphorylation dynamics in signaling networks.
  34. (1997). Glucoseinduced insulin secretion in INS-1 cells depends on factors present in fetal calf serum and rat islet-conditioned medium.
  35. (1996). ICA 512, an autoantigen of type I diabetes, is an intrinsic membrane protein of neurosecretory granules.
  36. (2005). Identification and biochemical analysis of Slac2-c/MyRIP as a Rab27A-, myosin Va/VIIa-, and actin-binding protein.
  37. (2004). Imaging Glucose-Regulated Insulin Secretion and Gene Expression in Single Islet betaCells: Control by AMP-Activated Protein Kinase.
  38. (2006). In-gel digestion for mass spectrometric characterization of proteins and proteomes.
  39. (1998). Interaction of muscle and brain sodium channels with multiple members of the syntrophin family of dystrophin-associated proteins.
  40. (1999). Interaction of neuronal nitric-oxide synthase with alpha1-syntrophin in rat brain.
  41. (1996). Interaction of nitric oxide synthase with the postsynaptic density protein PSD-95 and alpha1-syntrophin mediated by PDZ domains.
  42. (1999). Interactions between beta 2-syntrophin and a family of microtubule-associated serine/threonine kinases.
  43. (2003). Involvement of the Rab27 binding protein Slac2c/MyRIP in insulin exocytosis.
  44. (1994). Isolation, sequence and expression of a novel mouse brain cDNA, mIA-2, and its relatedness to members of the protein tyrosine phosphatase family.
  45. Kuroda TS (2002a) Slac2-c (synaptotagmin-like protein homologue lacking C2 domains-c), a novel linker protein that interacts with Rab27, myosin Va/VIIa, and actin.
  46. (2000). Membrane trafficking, organelle transport, and the cytoskeleton.
  47. (1994). Molecular cloning and identification of a receptor-type protein tyrosine phosphatase, IA-2, from human insulinoma.
  48. (1997). Motors and membrane traffic.
  49. (1995). Mouse alpha 1- and beta 2-syntrophin gene structure, chromosome localization, and homology with a discs large domain.
  50. (2008). Moving EM: the Rapid Transfer System as a new tool for correlative light and electron microscopy and high throughput for high-pressure freezing.
  51. (2005). Myosin 5a controls insulin granule recruitment during late-phase secretion.
  52. (2003). Myosin Va facilitates the distribution of secretory granules in the F-actin rich cortex of PC12 cells.
  53. (2005). Myosin Va transports dense core secretory vesicles in pancreatic MIN6 beta-cells.
  54. (2004). Nuclear translocation of an ICA512 cytosolic fragment couples granule exocytosis and insulin expression in {beta}-cells.
  55. (1988). Peripheral actin filaments control calcium-mediated catecholamine release from streptolysin-O-permeabilized chromaffin cells.
  56. (2006). Quantitative proteomics identifies Gemin5, a scaffolding protein involved in ribonucleoprotein assembly, as a novel partner for eukaryotic initiation factor 4E.
  57. (2005). Rab conversion as a mechanism of progression from early to late endosomes.
  58. (2003). Rab27A and its effector MyRIP link secretory granules to F-actin and control their motion towards release sites.
  59. (1999). Regulation of exocytosis by cyclin-dependent kinase 5 via phosphorylation of Munc18.
  60. (2008). Regulation of insulin granule turnover in pancreatic beta-cells by cleaved ICA512.
  61. (2004). Regulation of synaptojanin 1 by cyclin-dependent kinase 5 at synapses.
  62. (2000). Role of actin cortex in the subplasmalemmal transport of secretory granules in PC-12 cells.
  63. (2003). Secretory granules: and the last shall be first.
  64. (1996). Sitespecific phosphorylation of synapsin I by mitogen-activated protein kinase and Cdk5 and its effects on physiological functions.
  65. (2004). Slac2-a/melanophilin contains multiple PEST-like sequences that are highly sensitive to proteolysis.
  66. (1995). Syntrophin binds to an alternatively spliced exon of dystrophin.
  67. (2001). Syntrophin-dependent expression and localization of Aquaporin-4 water channel protein.
  68. (2002). Syntrophins and dystrobrevins: defining the dystrophin scaffold at synapses.
  69. (2003). The actin-binding domain of Slac2-a/ melanophilin is required for melanosome distribution in melanocytes.
  70. (2003). The cytoskeleton, cellular motility and the reductionist agenda.
  71. (1998). the exocytic pathway of MDCK cells.
  72. (2000). The receptor tyrosine phosphatase-like protein ICA512 binds the PDZ domains of beta2-syntrophin and nNOS in pancreatic beta-cells.
  73. (2006). The role of myosin Va in secretory granule trafficking and exocytosis.
  74. (1996). The three human syntrophin genes are expressed in diverse tissues, have distinct chromosomal locations, and each bind to dystrophin and its relatives.
  75. (2001). Total internal reflection fluorescence microscopy in cell biology.
  76. (2000). Tracking chromaffin granules on their way through the actin cortex.