Fan Yeung

University of Colorado at Boulder, Boulder, Colorado, United States

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Publications (17)93.19 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Sirtuins are class III deacetylases that regulate many essential processes, including cellular stress, genome stability and metabolism. Although these NAD(+)-dependent deacetylases control adaptive cellular responses, identification of sirtuin-regulated signaling targets remain under-studied. Here, we demonstrate that acetylation of the mitogen-activated protein kinase kinase-1 (MEK1) stimulates its kinase activity, and that acetylated MEK1 is under the regulatory control of the sirtuin family members SIRT1 and SIRT2. Treatment of cells with sirtuin inhibitors, or siRNA knockdown of SIRT1 or SIRT2 proteins, increases MEK1 acetylation and subsequent phosphorylation of the extracellular signal-regulated kinase. Generation of an acetyl-specific MEK1 antibody demonstrates that endogenous acetylated MEK1 is extensively enriched in the nucleus following epidermal growth factor stimulation. An acetyl-mimic of MEK1 increases inappropriate growth properties, suggesting that acetylation of MEK1 has oncogenic potential.Oncogene advance online publication, 31 March 2014; doi:10.1038/onc.2014.39.
    Oncogene 03/2014; · 8.56 Impact Factor
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    ABSTRACT: Calcium fluctuations and cardiac hypertrophy occur during pregnancy but the role of the well-studied calcium-activated phosphatase, calcineurin, has not been studied in this setting. The purpose of this study was to determine whether calcineurin signaling is required for cardiac remodeling during pregnancy in mice. We first examined calcineurin expression in the heart of mice during pregnancy. We found both calcineurin levels and activity were significantly increased in early-pregnancy and decreased in late-pregnancy. Since progesterone levels start to rise in early-pregnancy, we investigated whether progesterone alone was sufficient to modulate calcineurin levels in vivo. After implantation of progesterone pellets in non-pregnant female mice, cardiac mass increased, while cardiac function was maintained. In addition, calcineurin levels increased, which is also consistent with early-pregnancy. In order to determine whether these effects were occurring in the cardiac myocytes, we treated neonatal rat ventricular myocytes (NRVMs) with pregnancy-associated sex hormones. We found that progesterone treatment, but not estradiol, increased calcineurin levels. To obtain a functional readout of increased calcineurin activity, we measured the activity of the transcription factor NFAT, a downstream target of calcineurin. Progesterone treatment significantly increased NFAT activity in NRVMs, and this was blocked by the calcineurin inhibitor cyclosporine A (CsA), showing that the progesterone-mediated increase in NFAT activity requires calcineurin activity. Importantly, CsA treatment of mice completely blocked pregnancy-induced cardiac hypertrophy. Our results show that calcineurin is required for pregnancy-induced cardiac hypertrophy, and that calcineurin activity in early-pregnancy is due at least in part to increased progesterone.
    Cardiovascular Research 08/2013; · 5.81 Impact Factor
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    ABSTRACT: The sarcomeric myosin gene, Myh7b, encodes an intronic microRNA, miR-499, which regulates cardiac and skeletal muscle biology, yet little is known about its transcriptional regulation. To identify the transcription factors involved in regulating Myh7b/miR-499 gene expression, we have mapped the transcriptional start sites and identified an upstream 6.2 kb region of the mouse Myh7b gene whose activity mimics the expression pattern of the endogenous Myh7b gene both in vitro and in vivo. Through promoter deletion analysis, we have mapped a distal E-box element and a proximal Ikaros site that are essential for Myh7b promoter activity in muscle cells. We show that the myogenic regulatory factors, MyoD, Myf5 and Myogenin, bind to the E-box, while a lymphoid transcription factor, Ikaros 4 (Eos), binds to the Ikaros motif. Further, we show that through physical interaction, MyoD and Eos form an active transcriptional complex on the chromatin to regulate the expression of the endogenous Myh7b/miR-499 gene in muscle cells. We also provide the first evidence that Eos can regulate expression of additional myosin genes (Myosin 1 and β-Myosin) via the miR-499/Sox6 pathway. Therefore, our results indicate a novel role for Eos in the regulation of the myofiber gene program.
    Nucleic Acids Research 05/2012; 40(15):7303-18. · 8.81 Impact Factor
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    ABSTRACT: Although the signaling pathways underlying exercise-induced cardiac adaptation have been extensively studied, little is known about the molecular mechanisms that result in the response of the heart to pregnancy. The objective of this study was to define the morphological, functional, and gene expression patterns that define the hearts of pregnant mice, and to identify the signaling pathways that mediate this response. Mice were divided into three groups: nonpregnant diestrus control, midpregnancy, and late pregnancy. Both time points of pregnancy were associated with significant cardiac hypertrophy. The prosurvival signaling cascades of Akt and ERK1/2 were activated in the hearts of pregnant mice, while the stress kinase, p38, was decreased. Given the activation of Akt in pregnancy and its known role in cardiac hypertrophy, the hypertrophic response to pregnancy was tested in mice expressing a cardiac-specific activated (myristoylated) form of Akt (myrAkt) or a cardiac-specific constitutively active (antipathologic hypertrophic) form of its downstream target, glycogen synthase kinase 3β (caGSK3β). The pregnancy-induced hypertrophic responses of hearts from these mice were significantly attenuated. Finally, we tested whether pregnancy-associated sex hormones could induce hypertrophy and alter signaling pathways in isolated neonatal rat ventricular myocytes (NRVMs). In fact, progesterone, but not estradiol treatment increased NRVM cell size via phosphorylation of ERK1/2. Inhibition of MEK1 effectively blocked progesterone-induced cellular hypertrophy. Taken together, our study demonstrates that pregnancy-induced cardiac hypertrophy is mediated by activation of Akt and ERK1/2 pathways.
    Journal of Applied Physiology 02/2012; 112(9):1564-75. · 3.48 Impact Factor
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    ABSTRACT: To investigate the effect of bone environment on cellular proliferation, mature prostate-specific antigen (PSA) production and secretion, and PSA transcriptional regulation of prostate cancer cells. Androgen-independent C4-2 prostate cancer cells were co-cultured with various osteoblastic cells in a transwell system. Proliferation was measured via cell counting and MTT assay. Lactate and PSA were determined in the conditioned media (CM). Transcriptional activity of the full-length PSA promoter (6.1 kilobases) and of 3 deletion constructs was determined via luciferase reporter assay upon exposure to CM from various osteoblastic cell lines. Osteoblastic bone cells and CM, but not control cells (fibroblast) or CM, reproducibly stimulated the proliferation of C4-2 cells. The co-culture system, PSA production by C4-2 cells transiently decreased when in co-culture with osteoblastic, but not with control cells. After abundant prostate cell proliferation, the secreted PSA levels rose exponentially. Addition of CM from osteoblastic cells, but not control cells, consistently decreased (about 3-fold) the transcriptional activity of the PSA promoter in C4-2 cells. Deletion construct analysis of the PSA promoter revealed that the transcriptional down-regulation is dually controlled by elements close to the TATA and upstream androgen responsive (ARE(III)) components. The osteoblastic environment stimulates prostate cancer cell proliferation but reduces PSA production initially. The mechanism of PSA down-regulation is transcriptional, most likely in response to soluble factors present in the osteoblastic bone stromal cell CM. Transcriptional down-regulation appears to be mediated by elements near both the TATA box and the ARE(III) component.
    Urologic Oncology 11/2011; 29(6):802-8. · 3.65 Impact Factor
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    ABSTRACT: Nuclear factor-kappaB (NF-kappaB) is a dynamic transcription factor that regulates important biological processes involved in cancer initiation and progression. Identifying regulators that control the half-life of NF-kappaB is important to understanding molecular processes that control the duration of transcriptional responses. In this study we identify copine-I, a calcium phospholipid-binding protein, as a novel repressor that physically interacts with p65 to inhibit NF-kappaB transcription. Knockdown of copine-I by siRNA increases tumor necrosis factor alpha-stimulated NF-kappaB transcription, while copine-I expression blocks endogenous transcription. Copine-I abolishes NF-kappaB transcription by inducing endoprotease processing of the N-terminus of p65, a process antagonized by IkappaB alpha. Copine-I stimulates endoproteolysis of p65 within a conserved region that is required for base-specific contact with DNA. p65 proteins lacking the N-terminus fail to bind to DNA and act as dominant-negative molecules that inhibit NF-kappaB transcription. Our work provides evidence that copine-I regulates the half-life of NF-kappaB transcriptional responses through a novel mechanism that involves endoproteolysis of the p65 protein.
    Oncogene 07/2008; 27(25):3516-26. · 8.56 Impact Factor
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    ABSTRACT: Numerous mouse models of prostate carcinogenesis have been developed, but hitherto there has been no model in which the prostate gland could be imaged in live animals. The transgenic model generated here targeted mouse prostate gland using a firefly luciferase enzyme under the control of a small but highly active and specific supra prostate-specific antigen (sPSA) promoter. We evaluated postnatal prostate development, involution and androgen-induced restoration of prostate growth in adult transgenic mice using bioluminescence imaging. Results of our study showed that: (i) the prostate gland of male offspring did not yield a significant bioluminescence signal until after sexual maturity. Luciferase was detected in the luminal epithelial cells of the ventral and dorsolateral lobes of the prostate gland and caput epididymis, with little or no activity in 18 other organs evaluated. (ii) While a constant high level of bioluminescence was detected in the mouse prostate from 5 to 35 weeks of age, a slight drop in bioluminescence was detected at 36 to 54 weeks. (iii) Upon castration, the luciferase activity signal associated with mouse prostate detected by a cooled charge-coupled device camera was dramatically reduced. This signal could be rapidly restored to pre-castration levels after androgen administration. Androgen-induced luciferase activity subsided to nearly basal levels 5 days following the last injection. These data demonstrate that a bioluminescent mouse model with luciferase activity restricted to the prostate gland under the control of a (sPSA) promoter can be used on a real-time basis in live animals to investigate the development and responsiveness of the prostate gland to exogenously administered androgen. This model can be extended to detect the responsiveness of the prostate gland to therapy and used as a founder strain to visualize tumors in hosts with different genetic backgrounds.
    Journal of Molecular Endocrinology 11/2005; 35(2):293-304. · 3.58 Impact Factor
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    ABSTRACT: Understanding how signaling cascades stimulate chromatin-remodeling events through derepression is one of the foremost questions in the transcription field. Here, we demonstrate that NF-kappaB transcription requires IKKalpha to phosphorylate SMRT on chromatin, stimulating the exchange of corepressor for coactivator complexes. IKKalpha-induced phosphorylation coincides with a loss of chromatin-associated SMRT and HDAC3 and with nuclear export of the SMRT corepressor, events required for expression of the NF-kappaB-regulated cIAP-2 and IL-8 genes. Although SMRT derepression corresponds with the recruitment of TBL1/TBLR1, this complex alone is insufficient to relieve repression. Using a nonphosphorylatable SMRT protein, we demonstrate that IKKalpha-induced phosphorylation is required to recruit 14-3-3epsilon and Ubc5 for SMRT derepression. Failure of IKKalpha to stimulate the removal of SMRT from chromatin inhibits the recruitment of NF-kappaB to promoters, blocking transcription and sensitizing cells to apoptosis. Our work provides evidence that IKKalpha orchestrates SMRT derepression, a prerequisite for NF-kappaB transcription and survival.
    Molecular Cell 11/2004; 16(2):245-55. · 15.28 Impact Factor
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    ABSTRACT: NF-kappaB is responsible for upregulating gene products that control cell survival. In this study, we demonstrate that SIRT1, a nicotinamide adenosine dinucleotide-dependent histone deacetylase, regulates the transcriptional activity of NF-kappaB. SIRT1, the mammalian ortholog of the yeast SIR2 (Silencing Information Regulator) and a member of the Sirtuin family, has been implicated in modulating transcriptional silencing and cell survival. SIRT1 physically interacts with the RelA/p65 subunit of NF-kappaB and inhibits transcription by deacetylating RelA/p65 at lysine 310. Treatment of cells with resveratrol, a small-molecule agonist of Sirtuin activity, potentiates chromatin-associated SIRT1 protein on the cIAP-2 promoter region, an effect that correlates with a loss of NF-kappaB-regulated gene expression and sensitization of cells to TNFalpha-induced apoptosis. While SIRT1 is capable of protecting cells from p53-induced apoptosis, our work provides evidence that SIRT1 activity augments apoptosis in response to TNFalpha by the ability of the deacetylase to inhibit the transactivation potential of the RelA/p65 protein.
    The EMBO Journal 07/2004; 23(12):2369-80. · 9.82 Impact Factor
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    ABSTRACT: The NF-kappaB (nuclear factor-kappaB) transcription factors mediate activation of a large number of gene promoters containing diverse kappaB-site sequences. Here, PSA (prostate-specific antigen) was used as an AR (androgen receptor)-responsive gene to examine the underlying mechanism by which the NF-kappaB p65 transcription factor down-regulates the transcriptional activity of AR in cells. We observed that activation of NF-kappaB by TNFalpha (tumour necrosis factor alpha) inhibited both basal and androgen-stimulated PSA expression, and that this down-regulation occurred at the promoter level, as confirmed by the super-repressor IkappaBalpha (S32A/S36A), a dominant negative inhibitor of NF-kappaB. Using a linker-scanning mutagenesis approach, we identified a cis -element, designated XBE (X-factor-binding element), in the AREc (androgen response element enhancer core) of the PSA promoter, which negatively regulated several AR-responsive promoters, including that of PSA. When three copies of XBE in tandem were juxtaposed to GRE4 (glucocorticoid response element 4), a 4-6-fold reduction of inducible GRE4 activity was detected in three different cell lines, LNCaP, ARCaP-AR and PC3-AR. Bioinformatics and molecular biochemical studies indicated that XBE is a kappaB-like element that binds specifically to the NF-kappaB p65 subunit; consistent with these observations, only NF-kappaB p65, but not the NF-kappaB p50 subunit, was capable of inhibiting AR-mediated PSA promoter transactivation in LNCaP cells. In addition, our data also showed that AR binds to XBE, as well as to the kappaB consensus site, and that the transfection of AR inhibits the kappaB-responsive promoter in transient co-transfection assays. Collectively, these data indicate that cross-modulation between AR and NF-kappaB p65 transcription factors may occur by a novel mechanism involving binding to a common cis -DNA element.
    Biochemical Journal 05/2004; 379(Pt 2):421-31. · 4.65 Impact Factor
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    ABSTRACT: Prostate-specific antigen (PSA) is the most valuable marker for the evaluation of prostate cancer progression. The expression of PSA is controlled by androgen receptor (AR) through its binding to androgen-response elements (AREs). Several AREs have been identified within the 5.8-kb PSA promoter. The main activity of this 5.8-kb PSA promoter resides in a 455-bp enhancer core region located about 4 kb upstream of the TATA box. Our study suggests that in addition to the four AREs identified in the PSA enhancer core, another regulatory element (GAGATA), which is located at the region designated PSA3.1, also contributes to transcriptional regulation by androgens. Furthermore, electrophoretic mobility shift assay revealed that a putative transcriptional factor bound the GAGATA sequence in the PSA-producing prostate cancer cell. Further studies demonstrated that GAGATA factor preferentially bound the (G/C)(A/C/T)GATA sequence. The replacement of ATA with GGG in the GAGATA sequence completely eliminated the androgen-mediated transcriptional activity of the enhancer core. By using DNA-coupled magnetic beads and the Southwestern method, a 56-60-kDa protein was identified as the putative GAGATA binding factor. EMSA and Western blotting assay suggested that AR is not involved in androgen-mediated activation through PSA3.1. Therefore, we propose that binding of the GAGATA binding factor and AR to GAGATA and AREs, respectively, of the PSA enhancer core are required for the maximum transcriptional response to androgens.
    Journal of Biological Chemistry 09/2003; 278(34):32423-30. · 4.65 Impact Factor
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    ABSTRACT: Histone deacetylase (HDAC) inhibitors are emerging as a new class of anticancer agents for the treatment of solid and hematological malignancies. Although HDAC inhibitors induce cell death through an apoptotic process, little is known about the molecular events that control their effectiveness. In this study, we demonstrate that HDAC inhibitors are limited in their ability to induce apoptosis in non-small cell lung cancer (NSCLC) cell lines despite their ability to effectively inhibit deacetylase activity. Because the anti-apoptotic transcription factor NF-kappa B has been shown to be under the control of HDAC-mediated repression, we analyzed whether HDAC inhibitors activated NF-kappa B in NSCLC cells. HDAC inhibitors effectively stimulated endogenous NF-kappa B-dependent gene expression by up-regulating IL-8, Bcl-XL, and MMP-9 transcripts. The ability of HDAC inhibitors to increase NF-kappa B transcriptional activity was not associated with signaling events that stimulated nuclear translocation, but rather modulated the transactivation potential of the RelA/p65 subunit of NF-kappa B. The inhibition of HDAC activity was associated with the recruitment of the p300 transcriptional co-activator to chromatin in an Akt-dependent manner. Moreover, Akt directly phosphorylated p300 in vitro and was required for stimulating the transactivation potential of the co-activator following the addition of HDAC inhibitors. Selective inhibition of either the phosphoinositide 3-kinase/Akt pathway, or NF-kappa B itself blocked the ability of HDAC inhibitors to activate NF-kappa B and dramatically sensitized NSCLC cells to apoptosis following of the addition of HDAC inhibitors. Our study indicates that the ineffectiveness of HDAC inhibitors to induce apoptosis in NSCLC cancer cells is associated with the ability of these molecules to stimulate NF-kappa B-dependent transcription and cell survival.
    Journal of Biological Chemistry 06/2003; 278(21):18980-9. · 4.65 Impact Factor
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    ABSTRACT: The expression of prostate-specific membrane antigen (PSMA) and prostate-specific antigen (PSA), two well characterized marker proteins, remains highly active in the hormone refractory stage of prostate cancer. In this study, an artificial chimeric enhancer (PSES) composed of two modified regulatory elements controlling the expression of PSA and PSMA genes was tested for its promoter activity and tissue specificity using the reporter system. As a result, this novel PSES promoter remained silent in PSA- and PSMA-negative prostate and non-prostate cancer cell lines, but mediated high levels of luciferase in PSA- and PSMA-expressing prostate cancer cell lines in the presence and absence of androgen. To determine whether PSES could be used for in vivo gene therapy of prostate cancer, a recombinant adenovirus, Ad-PSES-luc, was constructed. Luciferase activity in prostate cancer cell lines mediated by Ad-PSES-luc was 400- to 1000-fold higher than in several other non-prostate cell lines, suggesting the high tissue-specificity of the PSES promoter in an adenoviral vector. Finally, recombinant virus Ad-PSES-luc was injected into mice to evaluate the tissue-discriminatory promoter activity in an experimental animal. Unlike Ad-CMV-luc, the luciferase activity from systemic injection of Ad-PSES-luc was fairly low in all major organs. However, when injected into prostate, Ad-PSES-luc drove high luciferase activity almost exclusively in prostate and not in other tissues. Our results demonstrated the potential use of PSES for the treatment of androgen-independent prostate cancer patients.
    Molecular Therapy 10/2002; 6(3):415-21. · 7.04 Impact Factor
  • Fan Yeung, Leland W K Chung
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    ABSTRACT: Prostate cancer is one of the leading causes of cancer death in Northern American men. The lethal phenotypes of human prostate cancer are characterized by progression to androgen-independence (Al) and a propensity to form osseous metastases. In approximately 80% of cases, prostate cancer colonizes bone and elicits a characteristic osteoblastic reaction. The bone metastases are initially sensitive to androgen deprivation treatments, but with time the cancer will eventually progress into an Al stage for which there is currently no effective treatment. Once initial hormonal therapy has failed, median survival of prostate cancer patients with bone metastases is less than 1 year (Tu et al. [2001] Lancet 357:336-341). Novel therapeutic and preventive strategies are needed to decrease morbidity and mortality of this disease.
    Journal of cellular biochemistry. Supplement 02/2002; 38:65-72.
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    ABSTRACT: Osteocalcin (OC) is a small (6 kDa) polypeptide whose expression was thought to be limited to mature osteoblasts. The discovery of OC expression in prostate cancer specimens led us to study the regulation of OC gene in androgen-independent metastatic human prostate PC3 cells. An 800-bp human OC (hOC) promoter-luciferase construct exhibited strong basal and vitamin D-induced activity in OC-positive human prostate and osteosarcoma cell lines. Through deletion analysis of the hOC promoter, the functional hierarchy of the cis-acting elements, OSE1, OSE2, and AP-1/VDRE, was established in PC3 cells (OSE1 > AP-1/VDRE > OSE2). By juxtaposing dimers of these 3 cis-elements, we produced a minimal hOC promoter capable of displaying high tissue specific activity in prostate cancer cells. Our study demonstrated three groups of transcription factors, Runx2, JunD/Fra-2, and Sp1, responsible for the high hOC promoter activity in PC3 cells by binding to the OSE2, AP-1/VDRE, and OSE1 elements, respectively. Among the three groups of transcription factors, the expression levels of Runx2 and Fra-2 are higher in the OC-positive PC3 cells and osteoblasts, compared with the OC-negative LNCaP cells. Interestingly, unlike the mouse OC promoter, the OSE1 site in hOC promoter is regulated by members of Sp1 family instead of the osteoblast-specific factor Osf1. The molecular basis for androgen-independent prostate cancer cells behaving like mature osteoblasts may be explained by the interplay and coordination of these transcription factors under the tight regulation of autocrine and paracrine mediators.
    Journal of Biological Chemistry 01/2002; 277(4):2468-76. · 4.65 Impact Factor
  • Journal of Urology - J UROL. 01/1999; 162(5).
  • Journal of Urology - J UROL. 01/1999; 162(5).