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Identification Of A Tripartite Basal Promoter Which Regulates Human Terminal Deoxynucleotidyl Transferase Gene Expression., D. Bhaumik, Baoli Yang, T. Trangas, J. Bartlett, M. Coleman, D. Sorscher Jul 2013

Identification Of A Tripartite Basal Promoter Which Regulates Human Terminal Deoxynucleotidyl Transferase Gene Expression., D. Bhaumik, Baoli Yang, T. Trangas, J. Bartlett, M. Coleman, D. Sorscher

Baoli Yang

In order to locate the promoter region of the human terminal deoxynucleotidyl transferase gene, serially truncated segments of the 5'-flanking region of the gene were cloned into a chloramphenicol acetyltransferase reporter vector. Transient transfection analyses of the terminal transferase-reporter gene constructs identified the basal promoter region within -34 to +40 base pairs relative to the transcription start site. Three promoter elements were defined in this region. The primary element is within 34 base pairs upstream of the transcription start site. The CAP site is 62 base pairs upstream of the translation start site. The secondary element involves sequences around the …


Loss Of Mll Phd Finger 3 Is Necessary For Mll-Enl-Induced Hematopoietic Stem Cell Immortalization, J. Chen, Donna Santillan, M. Koonce, W. Wei, R. Luo, M. Thirman, N. Zeleznik-Le, M. Diaz Jun 2013

Loss Of Mll Phd Finger 3 Is Necessary For Mll-Enl-Induced Hematopoietic Stem Cell Immortalization, J. Chen, Donna Santillan, M. Koonce, W. Wei, R. Luo, M. Thirman, N. Zeleznik-Le, M. Diaz

Donna A. Santillan

Reciprocal chromosomal translocations at the MLL gene locus result in expression of novel fusion proteins, such as MLL-ENL, associated with leukemia. The three PHD finger cassette, one of the highly conserved domains in MLL, is absent in all fusion proteins. This domain has been shown to interact with Cyp33, a cyclophilin which enhances the recruitment of histone deacetylases (HDAC) to the MLL repression domain and mediates HOX gene repression. Insertion of the third PHD finger of MLL into MLL-ENL allows the recruitment of Cyp33 and, subsequently, HDAC1 to the fusion protein. Furthermore, expression of the fusion protein with the PHD …


Expression And Localization Of Inducible And Endothelial Nitric Oxide Synthase In The Rat Ovary. Effects Of Gonadotropin Stimulation In Vivo, Bradley Van Voorhis, K. Moore, P. Strijbos, S. Nelson, S. Baylis, D. Grzybicki, C. Weiner Jun 2013

Expression And Localization Of Inducible And Endothelial Nitric Oxide Synthase In The Rat Ovary. Effects Of Gonadotropin Stimulation In Vivo, Bradley Van Voorhis, K. Moore, P. Strijbos, S. Nelson, S. Baylis, D. Grzybicki, C. Weiner

Bradley J Van Voorhis

Nitric oxide is reportedly involved in the regulation of several ovarian processes, yet the isoforms of nitric oxide synthase (NOS) expressed in the ovary are unknown. Our purpose was to identify and localize NOS isoenzymes in the rat ovary and to examine++ if mRNA expression of NOS isoenzymes change after gonadotropin stimulation. Using reverse transcriptase-PCR, we demonstrated that inducible (iNOS) and endothelial (eNOS), but not neuronal, NOS mRNAs are expressed in the ovary. In a gonadotropin-stimulated rat model, unstimulated ovaries had the highest levels of iNOS mRNA as quantified by ribonuclease protection assay. After gonadotropin injection, iNOS mRNA declined to …


Nitric Oxide: An Autocrine Regulator Of Human Granulosa-Luteal Cell Steroidogenesis, Bradley Van Voorhis, M. Dunn, G. Snyder, C. Weiner Jun 2013

Nitric Oxide: An Autocrine Regulator Of Human Granulosa-Luteal Cell Steroidogenesis, Bradley Van Voorhis, M. Dunn, G. Snyder, C. Weiner

Bradley J Van Voorhis

We investigated the presence of nitric oxide (NO) synthase in ovarian follicular cells obtained from women undergoing in vitro fertilization procedures. Endothelial NO synthase messenger RNA was demonstrated by polymerase chain reaction amplification of reverse transcribed RNA. NO synthase was localized to granulosa-luteal cells by immunocytochemistry, using a monoclonal antibody. Ovarian follicular cell NO synthase enzyme activity was confirmed by measuring the conversion of L-arginine to citrulline. To investigate the effect of NO on granulosa-luteal cell steroidogenesis, NO synthase inhibitors and NO donors were added to cell cultures. NG-Monomethyl-L-arginine and N-nitro-arginase methyl ester, selective inhibitors of NO synthase, significantly increased …


Cytoplasmic Metadherin (Mtdh) Provides Survival Advantage Under Conditions Of Stress By Acting As Rna-Binding Protein, Xiangbing Meng, Danlin Zhu, Shujie Yang, Xinjun Wang, Zhi Xiong, Yuping Zhang, Pavla Brachova, Kimberly Leslie May 2013

Cytoplasmic Metadherin (Mtdh) Provides Survival Advantage Under Conditions Of Stress By Acting As Rna-Binding Protein, Xiangbing Meng, Danlin Zhu, Shujie Yang, Xinjun Wang, Zhi Xiong, Yuping Zhang, Pavla Brachova, Kimberly Leslie

Xiangbing Meng

Overexpression of metadherin (MTDH) has been documented in many solid tumors and is implicated in metastasis and chemoresistance. MTDH has been detected at the plasma membrane as well as in the cytoplasm and nucleus, and the function of MTDH in these locales remains under investigation. In the nucleus, MTDH acts as a transcription co-factor to induce expression of chemoresistance-associated genes. However, MTDH is predominantly cytoplasmic in prostate tumors, and this localization correlates with poor prognosis. Herein, we used endometrial cancer cells as a model system to define a new role for MTDH in the cytoplasm. First, MTDH was primarily localized …


H19/Mir-675 Non-Coding Rna Expression Differentiates Among Cancers Of The Human Endometrium., Eric J. Devor, Jill N. Demik, Brandon M. Schickling, Michael J. Goodheart, Kimberly K. Leslie May 2013

H19/Mir-675 Non-Coding Rna Expression Differentiates Among Cancers Of The Human Endometrium., Eric J. Devor, Jill N. Demik, Brandon M. Schickling, Michael J. Goodheart, Kimberly K. Leslie

Michael J. Goodheart

H19 is a maternally expressed non-coding RNA located at chromosome 11p15.5 near the reciprocally imprinted insulin-like growth factor 2 (IGF2) gene. Though the function of H19 is unknown, it is transcribed during embryonic development after which transcription is absent in all but a few tissues including cardiac muscle, breast, ovary, uterus, and placenta. Linking H19, miR-675 and RB1 expression with serous tumors of the endometrium suggests that RB1 suppression may be a differentiating event in serous tumorigenesis.


H19/Mir-675 Non-Coding Rna Expression Differentiates Among Cancers Of The Human Endometrium., Eric J. Devor, Jill N. Demik, Brandon M. Schickling, Michael J. Goodheart, Kimberly K. Leslie May 2013

H19/Mir-675 Non-Coding Rna Expression Differentiates Among Cancers Of The Human Endometrium., Eric J. Devor, Jill N. Demik, Brandon M. Schickling, Michael J. Goodheart, Kimberly K. Leslie

Eric J Devor

H19 is a maternally expressed non-coding RNA located at chromosome 11p15.5 near the reciprocally imprinted insulin-like growth factor 2 (IGF2) gene. Though the function of H19 is unknown, it is transcribed during embryonic development after which transcription is absent in all but a few tissues including cardiac muscle, breast, ovary, uterus, and placenta. Linking H19, miR-675 and RB1 expression with serous tumors of the endometrium suggests that RB1 suppression may be a differentiating event in serous tumorigenesis.


Mrna Expression In Rabbit Experimental Aneurysms: A Study Using Gene Chip Microarrays, W. Mangrum, F. Farassati, R. Kadirvel, C. Kolbert, S. Raghavakaimal, Donghai Dai, Y. Ding, D. Grill, V. Khurana, D. Kallmes Apr 2013

Mrna Expression In Rabbit Experimental Aneurysms: A Study Using Gene Chip Microarrays, W. Mangrum, F. Farassati, R. Kadirvel, C. Kolbert, S. Raghavakaimal, Donghai Dai, Y. Ding, D. Grill, V. Khurana, D. Kallmes

Donghai Dai

BACKGROUND AND PURPOSE: The molecular characteristics of intracranial aneurysms are still poorly documented. A rabbit elastase aneurysm model has been helpful in the evaluation of devices and strategies involved in endovascular treatment of aneurysms. The goal of this project was to document the molecular changes, assessed by gene chip microarrays, associated with the creation of aneurysms in this model compared with the contralateral carotid artery. MATERIALS AND METHODS: A microarray of rabbit genes of interest was constructed using rabbit nucleotide sequences from GenBank. Elastase-induced saccular aneurysms were created at the origin of the right common carotid artery in 4 rabbits. …


Transfection Of C6 Glioma Cells With Glia Maturation Factor Upregulates Brain-Derived Neurotrophic Factor And Nerve Growth Factor: Trophic Effects And Protection Against Ethanol Toxicity In Cerebellar Granule Cells, N. Pantazis, A. Zaheer, Donghai Dai, S. Zaheer, S. Green, R. Lim Apr 2013

Transfection Of C6 Glioma Cells With Glia Maturation Factor Upregulates Brain-Derived Neurotrophic Factor And Nerve Growth Factor: Trophic Effects And Protection Against Ethanol Toxicity In Cerebellar Granule Cells, N. Pantazis, A. Zaheer, Donghai Dai, S. Zaheer, S. Green, R. Lim

Donghai Dai

Glial cells play active roles in neuronal survival, as well as neuroprotection against toxic insult. Recent studies suggest that the brain protein glia maturation factor (GMF) is involved in intracellular signaling in glia. This study investigated whether or not GMF plays a role in the survival-promoting and neuroprotective functions of glia. C6 glioma cells were transfected in vitro with GMF utilizing an adenovirus vector. The transfected cells overexpressed GMF intracellularly, but did not secrete the protein. The conditioned medium (CM) was obtained from the GMF-transfected cells (CM-GMF) and tested on primary neuronal cultures, consisting of cerebellar granule cells (CGC). The …


Relative Contribution Of Nedd4 And Nedd4-2 To Enac Regulation In Epithelia Determined By Rna Interference., Peter Snyder, Jennifer Steines, Diane Olson Mar 2013

Relative Contribution Of Nedd4 And Nedd4-2 To Enac Regulation In Epithelia Determined By Rna Interference., Peter Snyder, Jennifer Steines, Diane Olson

Jennifer C. Steines Wagemester

Epithelial Na+ transport is regulated in large part by mechanisms that control expression of the epithelial Na+ channel (ENaC) at the cell surface. Nedd4 and Nedd4-2 are candidates to control ENaC surface expression, but it is not known which of these proteins contributes to ENaC regulation in epithelia. To address this question, we used RNA interference to selectively reduce expression of Nedd4 or Nedd4-2. We found that endogenous Nedd4-2, but not Nedd4, negatively regulates ENaC in two epithelial cell lines (Fischer rat thyroid and H441); small interfering RNA (siRNA) against Nedd4-2 increased amiloride-sensitive Na+ current (compared with control siRNA), but …


Selective Down-Regulation Of Progesterone Receptor Isoform B In Poorly Differentiated Human Endometrial Cancer Cells: Implications For Unopposed Estrogen Action., N. Kumar, J. Richer, G. Owen, E. Litman, K. Horwitz, Kimberly Leslie Feb 2013

Selective Down-Regulation Of Progesterone Receptor Isoform B In Poorly Differentiated Human Endometrial Cancer Cells: Implications For Unopposed Estrogen Action., N. Kumar, J. Richer, G. Owen, E. Litman, K. Horwitz, Kimberly Leslie

Kimberly K. Leslie

The uterine endometrium responds to unopposed estrogen stimulation with rapid cell proliferation. Progesterone protects the endometrium against the hyperplastic effects of estradiol (E2) through progesterone receptors (PRs), of which two isoforms are expressed: human (h) PRA and PRB. hPRB has a longer NH2 terminus and may function differently from hPRA. Thus, the relative expression of hPRA:hPRB is likely to be important for the action of progesterone. We hypothesized that the hPRA:hPRB ratios may be abnormal in endometrial cancer, leading to a lack of normal progesterone protection against the growth-promoting effects of E2. To test this hypothesis, well-differentiated Ishikawa endometrial cancer …


Estrogen Receptors Are Present In Human Granulosa Cells., B. Hurst, M. Zilberstein, J. Chou, B. Litman, J. Stephens, Kimberly Leslie Feb 2013

Estrogen Receptors Are Present In Human Granulosa Cells., B. Hurst, M. Zilberstein, J. Chou, B. Litman, J. Stephens, Kimberly Leslie

Kimberly K. Leslie

Recent studies failed to detect estrogen receptors in primate follicles. This study was initiated to determine whether estrogen receptor (ER) messenger ribonucleic acid (mRNA) is present in human granulosa cells and, further, if functional ER proteins are present. To evaluate the presence of ER, RNA from human granulosa cells obtained at the time of oocyte retrieval for assisted reproduction was extracted, and complementary DNA synthesis was performed by the reverse transcriptase-polymerase chain reaction. Oligonucleotide primers were used to amplify basepairs 570-852 in the B- and C- domains of the ER mRNA. Southern blotting was performed and confirmed that the amplified …


Differential Gene Expression In The Amnion, Chorion, And Trophoblast Of The Human Placenta., C. Plouzek, Kimberly Leslie, J. Stephens, J. Chou Feb 2013

Differential Gene Expression In The Amnion, Chorion, And Trophoblast Of The Human Placenta., C. Plouzek, Kimberly Leslie, J. Stephens, J. Chou

Kimberly K. Leslie

Human chorionic gonadotrophin (hCG), placental alkaline phosphatase (PLAP), and pregnancy-specific glycoprotein (PSG) are three major proteins produced by the trophoblast of the human placenta. Immunocytochemical studies suggest that PSG and hCG are also present in the human amnion. In this study, we examined whether amniotic and chorionic membranes were capable of expressing trophoblastic-specific genes. As previously reported, trophoblasts express high levels of hCG beta, hCG alpha, PLAP, and PSG. Both amnion and chorion were found to express PLAP and hCG beta mRNA. However, the hCG alpha transcript was expressed only by the amnion, but not by the chorion in the …


The Role Of Oestrogen In Follicular Development., B. Hurst, Kimberly Leslie Feb 2013

The Role Of Oestrogen In Follicular Development., B. Hurst, Kimberly Leslie

Kimberly K. Leslie

Most of our knowledge of ovarian physiology is based upon studies that have demonstrated functional oestrogen receptors in the ovaries of lower animal species. The presence of oestrogen receptors in primate granulosa cells has been questioned by some investigators. However, we have found oestrogen receptor messenger RNA in human granulosa cells by reverse transcriptase-PCR assay. Furthermore, using immortalized granulosa cell lines transfected with a plasmid containing an oestrogen response element, a functional oestrogen receptor was confirmed. These experiments strongly support the hypothesis that human granulosa cells express biologically active oestrogen receptor.


Linkage Of Two Human Pregnancy-Specific Beta 1-Glycoprotein Genes: One Is Associated With Hydatidiform Mole., Kimberly Leslie, S. Watanabe, K. Lei, D. Chou, C. Plouzek, H. Deng, J. Torres, J. Chou Feb 2013

Linkage Of Two Human Pregnancy-Specific Beta 1-Glycoprotein Genes: One Is Associated With Hydatidiform Mole., Kimberly Leslie, S. Watanabe, K. Lei, D. Chou, C. Plouzek, H. Deng, J. Torres, J. Chou

Kimberly K. Leslie

A genomic clone containing two linked human pregnancy-specific beta 1-glycoprotein (PS beta G) genes has been isolated and characterized. The two genes are arranged in the same 5'----3' orientation; the 3' region (including the A2 and B-C exons) of the upstream gene, PSGGA, is linked to the 5' region (including the 5'/L and L/N exons) of PSGGB, the downstream gene. Depending upon the domains compared, PSGGA and PSGGB share 92-98% nucleotide and 86-95% amino acid sequence identity with PSG93, the most abundant PS beta G transcript. The 3' exon (B-C) of PSGGA contains four alternative splice sites and three polyadenylylation …


Cytoplasmic Metadherin (Mtdh) Provides Survival Advantage Under Conditions Of Stress By Acting As Rna-Binding Protein, Xiangbing Meng, Danlin Zhu, Shujie Yang, Xinjun Wang, Zhi Xiong, Yuping Zhang, Pavla Brachova, Kimberly Leslie Feb 2013

Cytoplasmic Metadherin (Mtdh) Provides Survival Advantage Under Conditions Of Stress By Acting As Rna-Binding Protein, Xiangbing Meng, Danlin Zhu, Shujie Yang, Xinjun Wang, Zhi Xiong, Yuping Zhang, Pavla Brachova, Kimberly Leslie

Kimberly K. Leslie

Overexpression of metadherin (MTDH) has been documented in many solid tumors and is implicated in metastasis and chemoresistance. MTDH has been detected at the plasma membrane as well as in the cytoplasm and nucleus, and the function of MTDH in these locales remains under investigation. In the nucleus, MTDH acts as a transcription co-factor to induce expression of chemoresistance-associated genes. However, MTDH is predominantly cytoplasmic in prostate tumors, and this localization correlates with poor prognosis. Herein, we used endometrial cancer cells as a model system to define a new role for MTDH in the cytoplasm. First, MTDH was primarily localized …


Progesterone Regulation Of Activating Protein-1 Transcriptional Activity: A Possible Mechanism Of Progesterone Inhibition Of Endometrial Cancer Cell Growth, Donghai Dai, E. Litman, E. Schonteich, Kimberly Leslie Feb 2013

Progesterone Regulation Of Activating Protein-1 Transcriptional Activity: A Possible Mechanism Of Progesterone Inhibition Of Endometrial Cancer Cell Growth, Donghai Dai, E. Litman, E. Schonteich, Kimberly Leslie

Kimberly K. Leslie

The uterine endometrium and cancers derived from it are classic models of hormone action: estrogen promotes growth and progesterone inhibits proliferation and results in differentiation. We have now identified a major pathway through which progesterone causes these growth-limiting effects. Ligand-bound progesterone receptors modulate the composition and transcriptional activity of members of the activating protein-1 (AP-1) family, and in particular, c-Jun. First, a dominant negative form of c-Jun inhibits the constitutive growth of Hec50co cells in a manner similar to the effects of progesterone through progesterone B receptors. Second, progesterone inhibits the transcriptional activity of the AP-1 complex in reporter gene …


H19/Mir-675 Non-Coding Rna Expression Differentiates Among Cancers Of The Human Endometrium., Eric J. Devor, Jill N. Demik, Brandon M. Schickling, Michael J. Goodheart, Kimberly K. Leslie Feb 2013

H19/Mir-675 Non-Coding Rna Expression Differentiates Among Cancers Of The Human Endometrium., Eric J. Devor, Jill N. Demik, Brandon M. Schickling, Michael J. Goodheart, Kimberly K. Leslie

Kimberly K. Leslie

H19 is a maternally expressed non-coding RNA located at chromosome 11p15.5 near the reciprocally imprinted insulin-like growth factor 2 (IGF2) gene. Though the function of H19 is unknown, it is transcribed during embryonic development after which transcription is absent in all but a few tissues including cardiac muscle, breast, ovary, uterus, and placenta. Linking H19, miR-675 and RB1 expression with serous tumors of the endometrium suggests that RB1 suppression may be a differentiating event in serous tumorigenesis.