title: Expression of a constitutively active prolactin receptor causes histone trimethylation of the p53 gene in breast cancer.
abstract: BACKGROUND
  Prolactin (PRL) is a pituitary polypeptide hormone characterized by multiple biological actions including stimulation of growth in the prostate and formation of secretory alveoli and stimulation of milk protein gene expression in the mammary gland. PRL exerts its effect by dimerizing its receptor (PRLR) on the plasma membrane and regulating gene expression through the JAK-Stat signal pathway. We have previously described a natural variant of the PRLR in which the S2 subdomain of the extracellular domain is missing (Delta S2). Delta S2 PRLRs are dimerized in the absence of PRL and have constitutive activity in the promotion of breast cancer cell growth. Enhancer of zeste homolog 2 (EZH2), as one of the histone-modifying enzymes, is a key factor regulating gene expression by epigenetic modification. We hypothesized that these constitutive activated Delta S2 PRLRs played a pathogenic role in breast cancer in part through alterations in the expression of EZH2 and the trimethylation of histone 3 on lysine 27 (H3K27Me3).
  
  
  METHODS
  In order to verify the clinical significance and to establish the link between Delta S2 PRLR expression and epigenetic change, EZH2, H3K27Me3, and Delta S2 PRLR were detected in both normal and cancerous human breast tissues. Also, overexpression of Delta S2 PRLR in breast epithelial cells was achieved by infection with adenovirus carrying the cDNA. Western blotting and chromatin immunoprecipitation (ChIP assay) and acid histone extraction were applied to detect the expression of EZH2 and the trimethylation of histone 3, respectively.
  
  
  RESULTS
  In breast tissue, higher EZH2 expression and higher H3K27Me3 were found associated with higher Delta S2 expression in breast cancer samples. In breast epithelial cells, overexpression of Delta S2 PRLR increased EZH2 methyltransferase mRNA and protein, induced EZH2 methyltransferase recruitment to chromatin, increased the trimethylation of H3K27Me3, and decreased the expression of p53 gene.
  
  
  CONCLUSIONS
  Delta S2 PRLR plays an important pathogenic role in breast cancer through epigenetic modification. Elevated expression of Delta S2 PRLR, achieved by alternate splicing of the pre-mRNA of the full-length form, is a new mechanism contributing to human breast cancer.
authors.affiliation: College of Medicine/Institute of Medical Sciences/Affiliated Hospital, Jishou University, Jishou, Hunan 416000, China. Email: naturet2002@yahoo.com.
authors.forename: Dunyong
authors.initials: D
authors.lastname: Tan
authors.affiliation: College of Medicine/Institute of Medical Sciences/Affiliated Hospital, Jishou University, Jishou, Hunan 416000, China.
authors.forename: Peizhi
authors.initials: P
authors.lastname: Tang
authors.affiliation: College of Medicine/Institute of Medical Sciences/Affiliated Hospital, Jishou University, Jishou, Hunan 416000, China.
authors.forename: Jianjun
authors.initials: J
authors.lastname: Huang
authors.affiliation: College of Medicine/Institute of Medical Sciences/Affiliated Hospital, Jishou University, Jishou, Hunan 416000, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.affiliation: College of Medicine/Institute of Medical Sciences/Affiliated Hospital, Jishou University, Jishou, Hunan 416000, China.
authors.forename: Weihua
authors.initials: W
authors.lastname: Zhou
authors.affiliation: Division of Biomedical Sciences, University of California Riverside, Riverside, CA 92521, USA.
authors.forename: Ameae M
authors.initials: AM
authors.lastname: Walker
mesh_terms: D001943:Breast Neoplasms / Q000378:metabolism*; D000071221:Enhancer of Zeste Homolog 2 Protein; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006657:Histones / Q000378:metabolism*; D006801:Humans; D061986:MCF-7 Cells; D063151:Polycomb Repressive Complex 2 / Q000378:metabolism*; D011981:Receptors, Prolactin / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
chemical_list: D006657:Histones; D011981:Receptors, Prolactin; D016159:Tumor Suppressor Protein p53; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D063151:Polycomb Repressive Complex 2