title: Deoxycholic acid activates protein kinase C and phospholipase C via increased Ca2+ entry at plasma membrane.
abstract: OBJECTIVE
  Secondary bile acids like deoxycholic acid (DCA) are well-established tumor promoters that may exert their pathologic actions by interfering with intracellular signaling cascades.
  
  
  METHODS
  We evaluated the effects of DCA on Ca2+ signaling in BHK-21 fibroblasts using fura-2 and mag-fura-2 to measure cytoplasmic and intraluminal internal stores [Ca2+], respectively. Furthermore, green fluorescent protein (GFP)-based probes were used to monitor time courses of phospholipase C (PLC) activation (pleckstrin-homology [PH]-PLCdelta-GFP), and translocation of protein kinase C (PKC) and a major PKC substrate, myristolated alanine-rich C-kinase substrate (MARCKS).
  
  
  RESULTS
  DCA (50-250 micromol/L) caused profound Ca2+ release from intracellular stores of intact or permeabilized cells. Correspondingly, DCA increased cytoplasmic Ca2+ to levels that were approximately 120% of those stimulated by Ca2+-mobilizing agonists in the presence of external Ca2+, and approximately 60% of control in Ca2+-free solutions. DCA also caused dramatic translocation of PH-PLCdelta-GFP, and conventional, Ca2+/diacylglycerol (DAG)-dependent isoforms of PKC (PKC-betaI and PKC-alpha), and MARCKS-GFP, but only in Ca2+-containing solutions. DCA had no effect on localization of a novel (PKCdelta) or an atypical (PKCzeta) PKC isoform.
  
  
  CONCLUSIONS
  Data are consistent with a model in which DCA directly induces both Ca2+ release from internal stores and persistent Ca2+ entry at the plasma membrane. The resulting microdomains of high Ca2+ levels beneath the plasma membrane appear to directly activate PLC, resulting in modest InsP 3 and DAG production. Furthermore, the increased Ca2+ entry stimulates vigorous recruitment of conventional PKC isoforms to the plasma membrane.
authors.affiliation: Boston VA Healthcare System and the Department of Surgery, Harvard Medical School, Brigham and Women's Hospital, 1400 VFW Parkway, West Roxbury, Massachusetts 02132, USA.
authors.forename: Bonnie W
authors.initials: BW
authors.lastname: Lau
authors.forename: Matilde
authors.initials: M
authors.lastname: Colella
authors.forename: Warren C
authors.initials: WC
authors.lastname: Ruder
authors.forename: Marianna
authors.initials: M
authors.lastname: Ranieri
authors.forename: Silvana
authors.initials: S
authors.lastname: Curci
authors.forename: Aldebaran M
authors.initials: AM
authors.lastname: Hofer
mesh_terms: D001692:Biological Transport / Q000187:drug effects; D018938:Caco-2 Cells; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D003840:Deoxycholic Acid / Q000494:pharmacology*; D004789:Enzyme Activation; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D011493:Protein Kinase C / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism*
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; C000619710:MARCKS protein, human; D008565:Membrane Proteins; D003840:Deoxycholic Acid; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D011493:Protein Kinase C; D010738:Type C Phospholipases; D002118:Calcium