Induction of PUMA-alpha and down-regulation of PUMA-beta expression is associated with benzo(a)pyrene-induced apoptosis in MCF-7 cells.
Benzo(a)pyrene (BP) forms benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE)-DNA adducts in human breast adenocarcinoma MCF-7 cells, leading to p53 protein induction and phosphorylation. Although BP-induced apoptosis in rodent cells is known, it is still unclear in human cells. Here we have analyzed the effects of BP on p53 related apoptotic proteins, cell cycle and cell death in MCF-7 cells. PUMA-protein (p53 up-regulated modulator of apoptosis) levels were changed after BP exposure so that PUMA-alpha protein was statistically significantly increased whereas PUMA-beta protein was statistically significantly decreased. PUMA-protein levels were also investigated in ZR-75-1 cells, where PUMA-alpha protein was statistically significantly increased. Cytochrome c, which is released from mitochondria during apoptosis to form the apoptosome, was increased in cytoplasmic fraction after BP exposure in MCF-7 cells. Increased apoptosis was also seen after 48 and 72 h BP exposure (2.5 and 5 microM). In addition, BP decreased dose dependently cell viability (2.5 and 5 microM) and increased ROS formation (1 and 10 microM). Our results suggest that PUMA-alpha protein is involved in BP-induced cell death most likely through a p53 dependent apoptotic pathway.
Tampio
Marjo
M
University of Kuopio, Department of Pharmacology and Toxicology, Yliopistonranta 1C, Kuopio, Finland. Marjo.Tampio@uku.fi
Markkanen
Piia
P
Puttonen
Katja A
KA
Hagelberg
Eveliina
E
Heikkinen
Hannu
H
Huhtinen
Kati
K
Loikkanen
Jarkko
J
Hirvonen
Maija-Riitta
MR
Vähäkangas
Kirsi H
KH
D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis*; D001564:Benzo(a)pyrene / Q000633:toxicity*; D045744:Cell Line, Tumor; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D015536:Down-Regulation; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D008564:Membrane Potentials / Q000187:drug effects; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis*; D017382:Reactive Oxygen Species / Q000378:metabolism; D015854:Up-Regulation
D051017:Apoptosis Regulatory Proteins; C438378:BBC3 protein, human; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins; D017382:Reactive Oxygen Species; D001564:Benzo(a)pyrene; D045304:Cytochromes c