Chromatin Modification of the Trefoil Factor 1 Gene in Human Breast Cancer Cells by the Ras/Mitogen-Activated Protein Kinase Pathway

Manitoba Institute of Cell Biology, Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Manitoba, Canada.
Cancer Research (Impact Factor: 9.33). 06/2006; 66(9):4610-6. DOI: 10.1158/0008-5472.CAN-05-4251
Source: PubMed


Histone H3 phosphorylation is a downstream response to activation of the Ras/mitogen-activated protein kinase (MAPK) pathway. This modification is thought to have a role in chromatin remodeling and in the initiation of gene transcription. In MCF-7 breast cancer cells, we observed that phosphorylated histone H3 (phospho-H3) at Ser(10) but not Ser(28) increased with phorbol ester (12-O-tetradecanoylphorbol-13-acetate, TPA) treatment. Although phosphorylated extracellular signal-regulated kinase 1/2 levels in these cells cultured under estradiol deplete and replete conditions displayed no change, a significant induction was observed after TPA treatment. Furthermore, whereas both estradiol and TPA increased trefoil factor 1 (TFF1) mRNA levels in these cells, only TPA-induced and not estradiol-induced TFF1 expression was inhibited by the H3 kinase mitogen and stress activated protein kinase (MSK) inhibitor H89 and MAPK kinase inhibitor UO126, showing the involvement of the Ras/MAPK following TPA induction. Mutation of the activator protein 1 (AP-1) binding site abrogated the TPA-induced transcriptional response of the luciferase reporter gene under the control of the TFF1 promoter, showing the requirement for the AP-1 site. In chromatin immunoprecipitation assays, estradiol treatment resulted in the association of the estrogen receptor-alpha (ERalpha) and acetylated H3 with the TFF1 promoter. The levels of phospho-H3 and MSK1 associated with the TFF1 promoter were moderately increased. In the presence of TPA, whereas ERalpha was not bound to the promoter, a strong association of acetylated and/or phospho-H3, MSK1, and c-Jun was observed. These results show that although both stimuli lead to TFF1 gene activation, estradiol and TPA exert their effects on TFF1 gene expression by different mechanisms.

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    • "Hormones (estradiol 11), growth factors such as insulin-like growth factor I (IGF-I), IGF-II, epidermal growth factor (EGF) or transforming growth factor (TGF) 11-13, phorbol esters 11, 14, plasminogen activator and Fos and Jun oncoproteins 11 have been reported to regulate the expression of the TFF1 through the activation of the gene promoter containing an estrogen-response element (ERE) and a TPA-response element (TRE) 15. The peptide was found to be overexpressed in approximately 50 % of primary breast carcinomas 16, mainly estrogen receptor alpha-positive (ERα+) ones. "
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    ABSTRACT: Background. A role of an estrogen-regulated, autocrine motogenic factor was assumed to be a major biological role of trefoil factor 1 (TFF1) in breast cancer. TFF1 is regarded as a predictive factor for positive response to endocrine therapy in breast cancer patients. The aim of our study was to examine TFF1 level distribution in breast carcinomas in order to distinguish estrogen-independent from estrogen-dependent TFF1 expression and to evaluate clinical usefulness of TFF1 status in early breast cancer during the first 3 years of follow-up. Methods. The study included 226 patients with primary operable invasive early breast carcinomas for whom an equal, a 3-year follow-up was conducted. TFF1 levels as well as estrogen receptor (ER) and progesterone receptor (PR) levels were measured in cytosolic extracts of tumor samples by immunoradiometric assay or by use of classical biochemical method, respectively. Non-parametric statistical tests were applied for data analyses. Results. Statistical analysis revealed that TFF1 levels were significantly higher in premenopausal patients (p=0.02), or in tumors with: lower histological grade (p<0.001), positive ER or PR status (p<0.001, in both cases). On the basis of TFF1 level distribution between ER-negative and ER-positive postmenopausal patients with tumors of different histological grade, 14 ng/mg was set as the cut-off value to distinguish estrogen-independent from estrogen-dependent TFF1 expression in breast cancer. Depending on menopausal and PR status, positive TFF1 status identified patients at opposite risk for relapse among ER-positive patients with grade II tumors. Among ER- and PR-positive premenopausal patients with grade II tumors, TFF1 status alone identified patients at opposite risk for relapse. Conclusions. Determination of TFF1 status might identify patients at different risk for relapse and help in making decision on administering adjuvant therapy for early breast cancer patients during the first 3 years of follow-up.
    Full-text · Article · May 2014 · International journal of medical sciences
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    • "Accordingly, we observed an increased HDAC2 or HDAC1 and SRSF1 co-occupancy on MCL1 exons 1 and 3, upon TPA induction (Figures 1D and Supplementary Figure S3A). Likewise, induction of the TFF1 gene in MCF7 cells by either estrogen or TPA (43,44) resulted in the co-occupancy of HDAC2 and SRSF1 along the body of the gene (Supplementary Figure S3C). Thus, the co-occupancy of HDAC1 or HDAC2 and SRSF1 along the body of transcribed genes was RNA-dependent, and occurred whether splicing was constitutive (FOSL1 and TFF1) or alternative (MCL1). "
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    ABSTRACT: Histone deacetylases (HDACs) and lysine acetyltransferases (KATs) catalyze dynamic histone acetylation at regulatory and coding regions of transcribed genes. Highly phosphorylated HDAC2 is recruited within corepressor complexes to regulatory regions, while the nonphosphorylated form is associated with the gene body. In this study, we characterized the nonphosphorylated HDAC2 complexes recruited to the transcribed gene body and explored the function of HDAC-complex-mediated dynamic histone acetylation. HDAC1 and 2 were coimmunoprecipitated with several splicing factors, including serine/arginine-rich splicing factor 1 (SRSF1) which has roles in alternative splicing. The co-chromatin immunoprecipitation of HDAC1/2 and SRSF1 to the gene body was RNA-dependent. Inhibition of HDAC activity and knockdown of HDAC1, HDAC2 or SRSF1 showed that these proteins were involved in alternative splicing of MCL1. HDAC1/2 and KAT2B were associated with nascent pre-mRNA in general and with MCL1 pre-mRNA specifically. Inhibition of HDAC activity increased the occupancy of KAT2B and acetylation of H3 and H4 of the H3K4 methylated alternative MCL1 exon 2 nucleosome. Thus, nonphosphorylated HDAC1/2 is recruited to pre-mRNA by splicing factors to act at the RNA level with KAT2B and other KATs to catalyze dynamic histone acetylation of the MCL1 alternative exon and alter the splicing of MCL1 pre-mRNA.
    Full-text · Article · Nov 2013 · Nucleic Acids Research
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    • "The chromatin remodeling events taking place at the TFF1 UPE have been extensively studied in estrogen-driven induction of TFF1 [28]–[30], whereas phorbol ester induction of TFF1 is not as well understood. Previously, we reported that TPA addition to MCF-7 breast cancer cells cultured under estrogen-free serum starved conditions resulted in the recruitment of MSK1 to the TFF1 UPE, along with increased H3S10ph (but not H3S28ph) and H3 acetylation (H3K9acK14ac) levels [27]. However, the recruitment of MSK2 or the mechanistic consequences of H3S10 phosphorylation in the context of the TPA-induced TFF1 transcriptional activation have not been explored. "
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    ABSTRACT: Mitogen- and stress-activated protein kinases 1 and 2 (MSK1 and MSK2), activated downstream of the ERK- and p38-mitogen-activated protein kinase pathways are involved in cell survival, proliferation and differentiation. Following mitogenic or stress stimuli, they mediate the nucleosomal response, which includes phosphorylation of histone H3 at serine 10 (H3S10ph) coupled with transcriptional activation of immediate-early genes. While MSK1 and MSK2 are closely related, their relative roles may vary with cellular context and/or stimuli. However, our knowledge of MSK2 recruitment to immediate-early genes is limited, as research has primarily focused on MSK1. Here, we demonstrate that both MSK1 and MSK2, regulate the phorbol ester 12-O-tetradecanoylphorbol-13-acetate induced expression of the breast cancer marker gene, trefoil factor 1 (TFF1), by phosphorylating H3S10 at its 5' regulatory regions. The MSK-mediated phosphorylation of H3S10 promotes the recruitment of 14-3-3 isoforms and BRG1, the ATPase subunit of the BAF/PBAF remodeling complex, to the enhancer and upstream promoter elements of TFF1. The recruited chromatin remodeling activity leads to the RNA polymerase II carboxy-terminal domain phosphorylation at the TFF1 promoter, initiating TFF1 expression in MCF-7 breast cancer cells. Moreover, we show that MSK1 or MSK2 is recruited to TFF1 regulatory regions, but as components of different multiprotein complexes.
    Full-text · Article · May 2013 · PLoS ONE
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