Early to Late Endosome Trafficking Controls Secretion and Zymogen Activation in Rodent and Human Pancreatic Acinar Cells
| Autor: | Diana D. H. Thomas, Michelle M. Cooley, Scott W. Messenger, Fred S. Gorelick, Michelle A. Falkowski, Luis A. Fernandez, Guy E. Groblewski, Benjamin K. August, Elaina K. Jones |
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| Rok vydání: | 2015 |
| Předmět: |
WT
wild type AP adaptor protein ZG zymogen granule CCK cholecystokinin DMSO dimethyl sulfoxide BFA brefeldin A PIKFYVE chemistry.chemical_compound PIKfyve phosphatidylinositol 3-phosphate 5-kinase GTP guanosine triphosphate Endosome Trypsin Trypsinogen activation Late endosome Original Research GDP guanosine diphosphate LY294002 2-morpholin-4-yl-8-phenylchromen-4-one LDH lactate dehydrogenase Gastroenterology Cell biology PI(3)P phosphatidylinositol 3-phosphate RE recycling endosome medicine.medical_specialty LAMP1 lysosome-associated membrane protein Phosphatidylinositol 3 5-bisphosphate Biology PI phosphatidylinositol Exocytosis MRP minor-regulated pathway VAMP8 vesicle-associated membrane protein 8 Internal medicine medicine Secretion lcsh:RC799-869 PI(3 5)P2 phosphatidylinositol-3 5-bisphosphate Hepatology TGN trans-Golgi network Phosphatidylinositol 3-phosphate DMEM Dulbecco’s minimal essential medium PIKfyve YM201636 6-amino-N-[3-(4-morpholin-4-ylpyrido[2 3]furo[2 4-b]pyrimidin-2-yl)phenyl]pyridine-3-carboxamide Vps34 phosphatylinositol 3-kinase Endocrinology EE early endosome Pancreatitis chemistry Zymogen activation CLP constitutive-like pathway GFP green fluorescent protein lcsh:Diseases of the digestive system. Gastroenterology LE late endosome HA hemagglutinin |
| Zdroj: | Cellular and Molecular Gastroenterology and Hepatology, Vol 1, Iss 6, Pp 695-709 (2015) Cellular and Molecular Gastroenterology and Hepatology |
| ISSN: | 2352-345X |
| DOI: | 10.1016/j.jcmgh.2015.08.002 |
| Popis: | Background & Aims: Pancreatic acinar cells have an expanded apical endosomal system, the physiologic and pathophysiologic significance of which is still emerging. Phosphatidylinositol-3,5-bisphosphate [PI(3,5)P2] is an essential phospholipid generated by phosphatidylinositol 3-phosphate 5-kinase (PIKfyve), which phosphorylates phosphatidylinositol-3-phosphate (PI3P). PI(3,5)P2 is necessary for maturation of early endosomes (EE) to late endosomes (LE). Inhibition of EE to LE trafficking enhances anterograde endosomal trafficking and secretion at the plasma membrane by default through a recycling endosome (RE) intermediate. We assessed the effects of modulating PIKfyve activity on apical trafficking and pancreatitis responses in pancreatic acinar cells. Methods: Inhibition of EE to LE trafficking was achieved using pharmacologic inhibitors of PIKfyve, expression of dominant negative PIKfyve K1877E, or constitutively active Rab5-GTP Q79L. Anterograde endosomal trafficking was manipulated by expression of constitutively active and dominant negative Rab11a mutants. The effects of these agents on secretion, endolysosomal exocytosis of lysosome associated membrane protein (LAMP1), and trypsinogen activation in response to supramaximal cholecystokinin (CCK-8), bile acids, and cigarette toxin was determined. Results: PIKfyve inhibition increased basal and stimulated secretion. Adenoviral overexpression of PIKfyve decreased secretion leading to cellular death. Expression of Rab5-GTP Q79L or Rab11a-GTP Q70L enhanced secretion. Conversely, dominant-negative Rab11a-GDP S25N reduced secretion. High-dose CCK inhibited endolysosomal exocytosis that was reversed by PIKfyve inhibition. PIKfyve inhibition blocked intracellular trypsin accumulation and cellular damage responses to supramaximal CCK-8, tobacco toxin, and bile salts in both rodent and human acini. Conclusions: These data demonstrate that EE-LE trafficking acutely controls acinar secretion and the intracellular activation of zymogens, leading to the pathogenicity of acute pancreatitis. Keywords: Endosome, Pancreatitis, PIKfyve, Trypsin |
| Databáze: | OpenAIRE |
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