Proteomic identification of <scp>E</scp>6<scp>AP</scp> as a molecular target of tamoxifen in <scp>MCF</scp>7 cells

Lochab, Savita; Pal, Pooja; Kanaujiya, Jitendra Kumar; Tripathi, Shashi B.; Kapoor, Isha; Bhatt, Madan Lal Brahma; Sanyal, Sabyasachi; Behre, Gerhard; Trivedi, Arun Kumar

doi:10.1002/pmic.201100572

Cited by 22 publications

(27 citation statements)

References 58 publications

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“…It was performed as previously described [23]. Briefly, 3T3-L1 preadipocytes pre-treated with MDI for 48 h were transiently transfected with expression plasmids for E6AP, E6AP-C843A and His-ubiquitin as indicated.…”

Section: Methodsmentioning

confidence: 99%

E3 Ubiquitin Ligase E6AP Negatively Regulates Adipogenesis by Downregulating Proadipogenic Factor C/EBPalpha

et al. 2013

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CCAAT/Enhancer Binding Protein Alpha (C/EBPα) is a key transcription factor involved in the adipocyte differentiation. Here for the first time we demonstrate that E6AP, an E3 ubiquitin ligase inhibits adipocyte differentiation in 3T3-L1 cells as revealed by reduced lipid staining with oil red. Knock down of E6AP in mouse 3T3L1 preadipocytes is sufficient to convert them to adipocytes independent of external hormonal induction. C/EBPα protein level is drastically increased in E6AP deficient 3T3L1 preadipocytes while inverse is observed when wild type E6AP is over expressed. We show that transient transfection of wild type E6AP downregulates C/EBPα protein expression in a dose dependent manner while catalytically inactive E6AP-C843A rather stabilizes it. In addition, wild type E6AP inhibits expression of proadipogenic genes while E6AP-C843A enhances them. More importantly, overexpression of E6AP-C843A in mesenchymal progenitor cells promotes accumulation of lipid droplets while there is drastically reduced lipid droplet formation when E6AP is over expressed. Taken together, our finding suggests that E6AP may negatively control adipogenesis by inhibiting C/EBPα expression by targeting it to ubiquitin-proteasome pathway for degradation.

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Section: Methodsmentioning

confidence: 99%

E3 Ubiquitin Ligase E6AP Negatively Regulates Adipogenesis by Downregulating Proadipogenic Factor C/EBPalpha

et al. 2013

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“…The ubiquitinprotein isopeptide ligase E6-associated protein (E6AP) associates with and promotes the degradation of ERα, which is inhibited by calmodulin. E6AP has been identified as a target for tamoxifen and is required for tamoxifen-mediated anti-breast cancer actions [162]. EFP can promote the ubiquitination and proteasome-dependent degradation of ERα in response to estrogen [154].…”

Section: Sumoylation Of Estrogen Receptormentioning

confidence: 99%

RETRACTED ARTICLE: Regulation of estrogen receptor signaling in breast carcinogenesis and breast cancer therapy

Zhou

Qiao

Shetty

et al. 2013

Cell. Mol. Life Sci.

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Estrogen and estrogen receptors (ERs) are critical regulators of breast epithelial cell proliferation, differentiation, and apoptosis. Compromised signaling vis-à-vis the estrogen receptor is believed © Springer Basel 2013 yow4@pitt.edu. ), which predicts for response to endocrine-based therapy; however, innate or acquired resistance to endocrinebased drugs remains a serious challenge. The complexity of regulation for estrogen signaling coupled with the crosstalk of other oncogenic signaling pathways is a reason for endocrine therapy resistance. Alternative strategies that target novel molecular mechanisms are necessary to overcome this current and urgent gap in therapy. A thorough analysis of estrogen-signaling regulation is critical. In this review article, we will summarize current insights into the regulation of estrogen signaling as related to breast carcinogenesis and breast cancer therapy. NIH Public Access

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“…Coimmunoprecipitation, immunocytochemistry, luciferase reporter assay, quantitative PCR (real-time) analysis, crystal violet staining, cell counting, morphological study, and Annexin V-PI staining were performed as described previously [38,39,41,42], and are also briefly mentioned in the supplementary details.…”

Section: Othersmentioning

confidence: 99%

Proteomic identification of Profilin1 as a corepressor of estrogen receptor alpha in MCF7 breast cancer cells

et al. 2013

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Nuclear receptor coregulators play an important role in the transcriptional regulation of nuclear receptors. In the present study, we aimed to identify estrogen receptor α (ERα) interacting proteins in Tamoxifen treated MCF7 cells. Using in vitro GST-pull down assay with ERα ligand-binding domain (ERα-LBD) and MS-based proteomics approach we identified Profilin1 as a novel ERα interacting protein. Profilin1 contains I/LXX/L/H/I amino acid signature motif required for corepressor interaction with ERα. We show that these two proteins physically interact with each other both in vitro as well as in vivo by GST-pull down and coimmunoprecipitation, respectively. We further show that these two proteins also colocalize together in the nucleus. Previous studies have reported reduced expression of Profilin1 in breast cancer; and here we found that Tamoxifen increases Profilin1 expression in MCF7 cells. Our data demonstrate that over expression of Profilin1 inhibits ERα-mediated transcriptional activation as well as its downstream target genes in ERα positive breast cancer cells MCF7. In addition, Profilin1 overexpression in MCF7 cells leads to inhibition of cell proliferation that apparently is due to enhanced apoptosis. In nutshell, these data indicate that MS-based proteomics approach identifies a novel ERα interacting protein Profilin1 that serves as a putative corepressor of ERα functions.

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Proteomic identification of E6AP as a molecular target of tamoxifen in MCF7 cells

Cited by 22 publications

References 58 publications

E3 Ubiquitin Ligase E6AP Negatively Regulates Adipogenesis by Downregulating Proadipogenic Factor C/EBPalpha

E3 Ubiquitin Ligase E6AP Negatively Regulates Adipogenesis by Downregulating Proadipogenic Factor C/EBPalpha

RETRACTED ARTICLE: Regulation of estrogen receptor signaling in breast carcinogenesis and breast cancer therapy

Proteomic identification of Profilin1 as a corepressor of estrogen receptor alpha in MCF7 breast cancer cells

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