title: Influence of cell cycle on responses of MCF-7 cells to benzo[a]pyrene.
abstract: BACKGROUND
  Benzo[a]pyrene (BaP) is a widespread environmental genotoxic carcinogen that damages DNA by forming adducts. This damage along with activation of the aryl hydrocarbon receptor (AHR) induces complex transcriptional responses in cells. To investigate whether human cells are more susceptible to BaP in a particular phase of the cell cycle, synchronised breast carcinoma MCF-7 cells were exposed to BaP. Cell cycle progression was analysed by flow cytometry, DNA adduct formation was assessed by 32P-postlabeling analysis, microarrays of 44K human genome-wide oligos and RT-PCR were used to detect gene expression (mRNA) changes and Western blotting was performed to determine the expression of some proteins, including cytochrome P450 (CYP) 1A1 and CYP1B1, which are involved in BaP metabolism.
  
  
  RESULTS
  Following BaP exposure, cells evaded G1 arrest and accumulated in S-phase. Higher levels of DNA damage occurred in S- and G2/M- compared with G0/G1-enriched cultures. Genes that were found to have altered expression included those involved in xenobiotic metabolism, apoptosis, cell cycle regulation and DNA repair. Gene ontology and pathway analysis showed the involvement of various signalling pathways in response to BaP exposure, such as the Catenin/Wnt pathway in G1, the ERK pathway in G1 and S, the Nrf2 pathway in S and G2/M and the Akt pathway in G2/M. An important finding was that higher levels of DNA damage in S- and G2/M-enriched cultures correlated with higher levels of CYP1A1 and CYP1B1 mRNA and proteins. Moreover, exposure of synchronised MCF-7 cells to BaP-7,8-diol-9,10-epoxide (BPDE), the ultimate carcinogenic metabolite of BaP, did not result in significant changes in DNA adduct levels at different phases of the cell cycle.
  
  
  CONCLUSIONS
  This study characterised the complex gene response to BaP in MCF-7 cells and revealed a strong correlation between the varying efficiency of BaP metabolism and DNA damage in different phases of the cell cycle. Our results suggest that growth kinetics within a target-cell population may be important determinants of susceptibility and response to a genotoxic agent.
authors.affiliation: Section of Molecular Carcinogenesis, Institute of Cancer Research, Brookes Lawley Building, Cotswold Road, Sutton, Surrey SM2 5NG, UK. hamza.hamouchene@icr.ac.uk
authors.forename: Hamza
authors.initials: H
authors.lastname: Hamouchene
authors.forename: Volker M
authors.initials: VM
authors.lastname: Arlt
authors.forename: Ian
authors.initials: I
authors.lastname: Giddings
authors.forename: David H
authors.initials: DH
authors.lastname: Phillips
mesh_terms: D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000378:metabolism; D001564:Benzo(a)pyrene / Q000633:toxicity*; D051177:Catenins / Q000235:genetics / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics / Q000378:metabolism; D065633:Cytochrome P-450 CYP1B1; D018736:DNA Adducts / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair / Q000235:genetics; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D053263:Gene Regulatory Networks; D006801:Humans; D051267:NF-E2-Related Factor 2 / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D025341:Principal Component Analysis; D015398:Signal Transduction / Q000235:genetics; D051153:Wnt Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D051177:Catenins; D018797:Cell Cycle Proteins; D018736:DNA Adducts; D051267:NF-E2-Related Factor 2; D051153:Wnt Proteins; D001564:Benzo(a)pyrene; D001189:Aryl Hydrocarbon Hydroxylases; C585496:CYP1B1 protein, human; D019363:Cytochrome P-450 CYP1A1; D065633:Cytochrome P-450 CYP1B1; D048049:Extracellular Signal-Regulated MAP Kinases