title: Role of intracellular calcium in proteasome inhibitor-induced endoplasmic reticulum stress, autophagy, and cell death.
abstract: OBJECTIVE
  Proteasome inhibition induces endoplasmic reticulum (ER) stress and compensatory autophagy to relieve ER stress. Disturbance of intracellular calcium homeostasis can lead to ER stress and alter the autophagy process. It has been suggested that inhibition of the proteasome disrupts intracellular calcium homeostasis. However, it is unknown if intracellular calcium affects proteasome inhibitor-induced ER stress and autophagy.
  
  
  METHODS
  Human colon cancer HCT116 Bax positive and negative cell lines were treated with MG132, a proteasome inhibitor. BAPTA-AM, a cell permeable free calcium chelator, was used to modulate intracellular calcium levels. Autophagy and cell death were determined by fluorescence microscopy and immunoblot analysis.
  
  
  RESULTS
  MG132 increased intracellular calcium levels in HCT116 cells, which was suppressed by BAPTA-AM. MG132 suppressed proteasome activity independent of Bax and intracellular calcium levels in HCT116 cells. BAPTA-AM inhibited MG132-induced cellular vacuolization and ER stress, but not apoptosis. MG132 induced autophagy with normal autophagosome-lysosome fusion. BAPTA-AM seemed not to affect autophagosome-lysosome fusion in MG132-treated cells but further enhanced MG132-induced LC3-II levels and GFP-LC3 puncta formation, which was likely via impaired lysosome function.
  
  
  CONCLUSIONS
  Blocking intracellular calcium by BAPTA-AM relieved MG132-induced ER stress, but it was unable to rescue MG132-induced apoptosis, which was likely due to impaired autophagic degradation.
authors.affiliation: Department of Pharmacology, Toxicology and Therapeutics, The University of Kansas Medical Center, MS 1018 3901 Rainbow Blvd., Kansas City, Kansas 66160, USA.
authors.forename: Jessica A
authors.initials: JA
authors.lastname: Williams
authors.forename: Yifeng
authors.initials: Y
authors.lastname: Hou
authors.forename: Hong-Min
authors.initials: HM
authors.lastname: Ni
authors.forename: Wen-Xing
authors.initials: WX
authors.lastname: Ding
mesh_terms: D001343:Autophagy / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D016923:Cell Death / Q000187:drug effects*; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects*; D045325:HCT116 Cells; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology*; D061988:Proteasome Inhibitors / Q000494:pharmacology*
chemical_list: D002614:Chelating Agents; D007976:Leupeptins; D061988:Proteasome Inhibitors; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; D002118:Calcium
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