title: Apoptosis induction of 2'-hydroxycinnamaldehyde as a proteasome inhibitor is associated with ER stress and mitochondrial perturbation in cancer cells.
abstract: 2'-Hydroxycinnamaldehyde (HCA), isolated from the stem bark of Cinnamomum cassia, and 2'-benzoyloxycinnamaldehyde (BCA), one of HCA derivatives, have antiproliferative activities on several human cancer cell lines. Our previous study suggested that reactive oxygen species (ROS) and caspase-3 are the major regulators of HCA-induced apoptosis. In the present study, we demonstrated a novel molecular target using in vitro pull-down assay by biotin-labeled HCA (biotin-HCA) in SW620 cells. We analyzed 11 differential spots of 2-dimensional gel prepared with pull-downed proteins by biotin-HCA. Among them, five spots were identified as proteasome subunits. An in vitro 26S proteasome function assay using specific fluorogenic substrates showed that HCA potently inhibits L3-like activity of the proteasome. In addition, HCA showed inhibitory action against chymotrypsin-like, trypsin-like, and PGPH-like activities. DNA microarray showed that HCA induced heat shock family and ER stress-responsive genes, which reflects the accumulation of misfolded proteins by proteasome inhibition. On western blot analysis, it was confirmed that HCA induces glucose-regulated protein, 78 kDa (GRP78) and some representative endoplasmic reticulum (ER) stress-responsive proteins. Furthermore, HCA treatment decreased mitochondrial membrane potential. The effect of HCA on cytochrome c and Bax translocation between cytosol and mitochondrial membrane was clarified using western blot analysis. These results suggest that HCA-induced apoptosis is associated with the inhibition of the proteasome activity that leads in turn to the increase of ER stress and mitochondrial perturbation.
authors.affiliation: Department of Dental Microbiology, School of Dentistry, Kyungpook National University, Daegu 700-412, Republic of Korea.
authors.forename: Su Hyung
authors.initials: SH
authors.lastname: Hong
authors.forename: Jina
authors.initials: J
authors.lastname: Kim
authors.forename: Jung-Min
authors.initials: JM
authors.lastname: Kim
authors.forename: So-Young
authors.initials: SY
authors.lastname: Lee
authors.forename: Dae-Seop
authors.initials: DS
authors.lastname: Shin
authors.forename: Kwang-Hee
authors.initials: KH
authors.lastname: Son
authors.forename: Dong Cho
authors.initials: DC
authors.lastname: Han
authors.forename: Young Kwan
authors.initials: YK
authors.lastname: Sung
authors.forename: Byoung-Mog
authors.initials: BM
authors.lastname: Kwon
mesh_terms: D036541:ADP-ribosyl Cyclase / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002934:Cinnamates / Q000032:analysis / Q000737:chemistry / Q000494:pharmacology*; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004721:Endoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism; D000091342:Endoplasmic Reticulum Chaperone BiP; D020869:Gene Expression Profiling; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008933:Mitochondrial Swelling / Q000187:drug effects*; D015394:Molecular Structure; D009369:Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D010711:Phosphatidate Phosphatase / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D061988:Proteasome Inhibitors; D021381:Protein Transport / Q000187:drug effects; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D051742:Transcription Factor CHOP / Q000235:genetics / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism
chemical_list: D002934:Cinnamates; D000091342:Endoplasmic Reticulum Chaperone BiP; C485932:HERPUD1 protein, human; C000717247:HSPA5 protein, human; D008565:Membrane Proteins; D061988:Proteasome Inhibitors; D051028:bcl-2-Associated X Protein; C117045:p-hydroxycinnamaldehyde; D051742:Transcription Factor CHOP; D045304:Cytochromes c; C496541:HMOX1 protein, human; D051547:Heme Oxygenase-1; C112948:lipid phosphate phosphatase; D010711:Phosphatidate Phosphatase; D036541:ADP-ribosyl Cyclase; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease