Thyroid hormone receptors (c-erbAs) and retinoic acid receptors (RARs) are members of a subgroup of closely related nuclear receptor proteins. c-erbAs and RAR can each activate certain response elements and both receptors contain a highly conserved domain embedded within the ligand binding region containing a series of """"""""leucine-zipper-like"""""""" hydrophobic heptad motifs. Functional studies suggest that the heptad repeat domain mediates homo- and heterodimeric interactions of c-erbA and RAR or interactions with other factors. Chick c-erbA-alpha and human RAR-alpha have been expressed in E. coli and purified to near homogeneity. These receptors bind ligand with appropriate affinity and form homo- and hetero-dimers on response elements which are permissive for dimerization. Dimer formation is enhanced by ligand suggesting that ligand mediates transcriptional activation by this mechanism. This application is a comprehensive proposal to define the functional domains involved in c-erbA, RAR, and related factors in transcriptional activation. For these studies we constructed a multifunctional bacterial/eucaryotic expression vector (pEXPRESS) which permits site directed mutagenesis and can be used to functionally analyze receptor in eucaryotic cells and to express receptor at high levels in E. coli. Gel shift studies using purified wild-type and mutant receptor proteins and a variety of native and synthetic response elements are proposed to elucidate the """"""""rules"""""""" which govern how these receptors recognize functional response elements. These studies will also define receptor domains critical for homo- and hetero-dimer formation, for cooperative interactions between receptors and other factors, and for dominant negative activities. Possible differences in element recognition by c-erbA subtypes (alphal and betal) will also be examined along with studies to understand differences between v-erbA and c-erbA and how c-erbA-alpha2 functions as a dominant negative regulator. These studies will be complemented by analyzing receptor mutants derived from patients with the thyroid hormone resistance syndrome. Functional studies in mammalian cells will be extended to in vitro transcription with the goal of defining the protein and DNA requirements for transcriptional enhancement by these receptors. Finally, the availability of mg amounts of purified E. coli expressed wild-type c-erbA and its DNA binding domain will allow circular dichroism, fluorescence, and ultraviolet-visible spectroscopy (both proteins), and nuclear magnetic resonance studies (DNA binding domain) to provide structural information to elucidate the functional and physical properties of these proteins at the molecular level.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
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Endocrinology Study Section (END)
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Margolis, Ronald N
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New York University
Schools of Medicine
New York
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Yang, Yawei J; Baltus, Andrew E; Mathew, Rebecca S et al. (2012) Microcephaly gene links trithorax and REST/NRSF to control neural stem cell proliferation and differentiation. Cell 151:1097-112
Yeung, Kay T; Das, Sharmistha; Zhang, Jin et al. (2011) A novel transcription complex that selectively modulates apoptosis of breast cancer cells through regulation of FASTKD2. Mol Cell Biol 31:2287-98
Vukelic, Sasa; Stojadinovic, Olivera; Pastar, Irena et al. (2010) Farnesyl pyrophosphate inhibits epithelialization and wound healing through the glucocorticoid receptor. J Biol Chem 285:1980-8
Goyanka, Ritu; Das, Sharmistha; Samuels, Herbert H et al. (2010) Nuclear receptor engineering based on novel structure activity relationships revealed by farnesyl pyrophosphate. Protein Eng Des Sel 23:809-15
Garapaty, Shivani; Xu, Chong-Feng; Trojer, Patrick et al. (2009) Identification and characterization of a novel nuclear protein complex involved in nuclear hormone receptor-mediated gene regulation. J Biol Chem 284:7542-52
Mahajan, Muktar A; Samuels, Herbert H (2008) Nuclear receptor coactivator/coregulator NCoA6(NRC) is a pleiotropic coregulator involved in transcription, cell survival, growth and development. Nucl Recept Signal 6:e002
Das, Sharmistha; Schapira, Matthieu; Tomic-Canic, Marjana et al. (2007) Farnesyl pyrophosphate is a novel transcriptional activator for a subset of nuclear hormone receptors. Mol Endocrinol 21:2672-86
Pevsner, Paul H; Naftolin, Frederick; Hillman, Dean E et al. (2007) Direct identification of proteins from T47D cells and murine brain tissue by matrix-assisted laser desorption/ionization post-source decay/collision-induced dissociation. Rapid Commun Mass Spectrom 21:429-36
Mahajan, Muktar A; Murray, Audrey; Levy, David et al. (2007) Nuclear receptor coregulator (NRC): mapping of the dimerization domain, activation of p53 and STAT-2, and identification of the activation domain AD2 necessary for nuclear receptor signaling. Mol Endocrinol 21:1822-34
Das, Sharmistha; Nwachukwu, Jerome C; Li, Dangsheng et al. (2007) The nuclear receptor interacting factor-3 transcriptional coregulator mediates rapid apoptosis in breast cancer cells through direct and bystander-mediated events. Cancer Res 67:1775-82

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