title: Regulation of Bcl-2 protein expression in human neuroblastoma SH-SY5Y cells: positive and negative effects of protein kinases C and A, respectively.
abstract: The regulatory mechanism of Bcl-2 protein expression was investigated in SH-SY5Y cells, the human neuroblastoma cell line that expresses natively Bcl-2 proteins. WHen the cells were treated with 12-O-tetradecanoylphorbol 13-acetate (TPA) or retinoic acid, the level of Bcl-2 protein was increased compared with the control. These effects were inhibited by pretreatment with a protein kinase C (PKC) inhibitor, staurosporine or calphostin C. The level of Bcl-2 protein was also increased by treatment with carbachol, a muscarinic acetylcholine receptor (mAChR) agonist, and the effect were also inhibited by pretreatment with staurosporine or calphostin C. An addition, a carbachol-induced increase in Bcl-2 protein levels and a transient elevation of [Ca2+]i were inhibited by pretreatment with 4-DAMP (4-diphenylacetoxy-N-methylpiperidine), an m3 mAChR antagonist. In contrast, the level of Bcl-2 protein was decreased by treatment with dibutyryl cAMP (diBu-cAMP), forskolin, or cholera toxin, and the effects of diBu-cAMP were inhibited by pretreatment with a protein kinase A (PKA) inhibitor, H-89. From these results, we suggest that the expression of Bcl-2 proteins is regulated by PKC and PKA in positive and negative manners, respectively, in SH-SY5Y cells. Furthermore, the nucleosomal DNA fragmentation induced by serum depletion for 4 h was observed in SH-SY5Y cells when the level of Bcl-2 protein was down-regulated by treatment with 1 mM diBu-cAMP for 3 days, although the DNA fragmentation by serum depletion for 4 h was not observed in nontreatment cells, indicating that Bcl-2 proteins whose expression is regulated by PKC and PKA play important roles in serum depletion-induced apoptosis.
authors.affiliation: Department of Pharmacology, Faculty of Pharmaceutical Chemistry, Hokkaido University, Sapporo, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Itano
authors.forename: A
authors.initials: A
authors.lastname: Ito
authors.forename: T
authors.initials: T
authors.lastname: Uehara
authors.forename: Y
authors.initials: Y
authors.lastname: Nomura
mesh_terms: D017209:Apoptosis / Q000502:physiology; D015152:Blotting, Northern; D015153:Blotting, Western; D003994:Bucladesine / Q000494:pharmacology; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology*; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D006801:Humans; D009447:Neuroblastoma; D011493:Protein Kinase C / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger / Q000032:analysis; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000201:enzymology
chemical_list: D016895:Culture Media, Serum-Free; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D003994:Bucladesine; D011505:Protein-Tyrosine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C