scholarly journals Adenovirus-directed Expression of Dominant Negative Estrogen Receptor Induces Apoptosis in Breast Cancer Cells and Regression of Tumors in Nude Mice

2001 ◽  
Vol 7 (11) ◽  
pp. 773-782 ◽  
Author(s):  
Eun Jig Lee ◽  
Monika Jakacka ◽  
W. Rachel Duan ◽  
Pei Yu Chien ◽  
Fred Martinson ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Estrogen Receptor ◽  
Cancer Cells ◽  
Nude Mice ◽  
Breast Cancer Cells ◽  
Dominant Negative

scholarly journals A Fusion Protein of the Estrogen Receptor (ER) and Nuclear Receptor Corepressor (NCoR) Strongly Inhibits Estrogen-Dependent Responses in Breast Cancer Cells

1999 ◽  
Vol 13 (12) ◽  
pp. 2122-2136 ◽  
Author(s):  
Pei-Yu Chien ◽  
Masafumi Ito ◽  
Youngkyu Park ◽  
Tetsuya Tagami ◽  
Barry D. Gehm ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Estrogen Receptor ◽  
Dna Binding ◽  
Fusion Protein ◽  
Cancer Cells ◽  
Nuclear Receptor ◽  
Fusion Proteins ◽  
Breast Cancer Cells ◽  
Dominant Negative ◽  
Dominant Negative Activity

Abstract Nuclear receptor corepressor (NCoR) mediates repression (silencing) of basal gene transcription by nuclear receptors for thyroid hormone and retinoic acid. The goal of this study was to create novel estrogen receptor (ER) mutants by fusing transferable repressor domains from the N-terminal region of NCoR to a functional ER fragment. Three chimeric NCoR-ER proteins were created and shown to lack transcriptional activity. These fusion proteins silenced basal transcription of the ERE2-tk-Luc reporter gene and inhibited the activity of cotransfected wild-type ER (wtER), indicating that they possess dominant negative activity. One of the fusion proteins (CDE-RD1), containing the ER DNA-binding and ligand-binding domains linked to the NCoR repressor domain (RD1), was selected for detailed examination. Its hormone affinity, intracellular localization, and level of expression in transfected cells were similar to wtER, and it bound to the estrogen response element (ERE) DNA in gel shift assays. Glutathione-S-transferase pull-down assays showed that CDE-RD1 retains the ability to bind to steroid receptor coactivator-1. Introduction of a DNA-binding domain mutation into the CDE-RD1 fusion protein eliminated silencing and dominant negative activity. Thus, the RD1 repressor domain prevents transcriptional activation despite the apparent ability of CDE-RD1 to bind DNA, ligand, and coactivators. Transcriptional silencing was incompletely reversed by trichostatin A, suggesting a histone deacetylase-independent mechanism for repression. CDE-RD1 inhibited ER-mediated transcription in T47D and MDA-MB-231 breast cancer cells and repressed the growth of T47D cells when delivered to the cells by a retroviral vector. These ER-NCoR fusion proteins provide a novel means for inhibiting ER-mediated cellular responses, and analogous strategies could be used to create dominant negative mutants of other transcription factors.

scholarly journals Functional role of TRPC6 and STIM2 in cytosolic and endoplasmic reticulum Ca2+ content in resting estrogen receptor-positive breast cancer cells

2020 ◽  
Vol 477 (17) ◽  
pp. 3183-3197
Author(s):  
Jose Sanchez-Collado ◽  
Jose J. Lopez ◽  
Lucia Gonzalez-Gutierrez ◽  
Carlos Cantonero ◽  
Isaac Jardin ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Endoplasmic Reticulum ◽  
Estrogen Receptor ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Dominant Negative ◽  
Cellular Functions ◽  
Mcf7 Cells ◽  
Membrane Expression ◽  
Estrogen Receptor Positive

TRPC6 forms non-selective cation channels activated by a variety of stimuli that are involved in a wide number of cellular functions. In estrogen receptor-positive (ER+) breast cancer cells, the store-operated Ca2+ entry has been reported to be dependent on STIM1, STIM2 and Orai3, with TRPC6 playing a key role in the activation of store-operated Ca2+ entry as well as in proliferation, migration and viability of breast cancer cells. We have used a combination of biotinylation, Ca2+ imaging as well as protein knockdown and overexpression of a dominant-negative TRPC6 mutant (TRPC6dn) to show that TRPC6 and STIM2 are required for the maintenance of cytosolic and endoplasmic reticulum Ca2+ content under resting conditions in ER+ breast cancer MCF7 cells. These cells exhibit a greater plasma membrane expression of TRPC6 under resting conditions than non-tumoral breast epithelial cells. Attenuation of STIM2, TRPC6 and Orai3, alone or in combination, results in impairment of resting cytosolic and endoplasmic reticulum Ca2+ homeostasis. Similar results were observed when cells were transfected with expression plasmid for TRPC6dn. TRPC6 co-immunoprecipitates with STIM2 in resting MCF7 cells, a process that is impaired by rises in cytosolic Ca2+ concentration. Impairment of TRPC6 function leads to abnormal Ca2+ homeostasis and endoplasmic reticulum stress, thus, suggesting that TRPC6 might be a potential target for the development of anti-tumoral therapies.

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