Transforming growth factor-beta inhibits aromatase gene transcription in human trophoblast cells via the Smad2 signaling
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Transforming growth factor-beta inhibits aromatase gene transcription in human trophoblast cells via the Smad2 signaling pathway Hong Zhou1,2, Guodong Fu1, Hui Yu1 and Chun Peng*1 Address: 1Department of Biology, York University, Toronto, Ontario, M3J 1P3, Canada and 2School of Life Science and Technology, University of Electronic Science and Technology of China, Chengdu, PR China Email: Hong Zhou - [email protected]; Guodong Fu - [email protected]; Hui Yu - [email protected]; Chun Peng* - [email protected] * Corresponding author
Published: 9 December 2009 Reproductive Biology and Endocrinology 2009, 7:146
doi:10.1186/1477-7827-7-146
Received: 26 October 2009 Accepted: 9 December 2009
This article is available from: http://www.rbej.com/content/7/1/146 © 2009 Zhou et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract Background: Transforming growth factor-beta (TGF-beta) is known to exert multiple regulatory functions in the human placenta, including inhibition of estrodial production. We have previously reported that TGF-beta1 decreased aromatase mRNA levels in human trophoblast cells. The objective of this study was to investigate the molecular mechanisms underlying the regulatory effect of TGF-beta1 on aromatase expression. Methods: To determine if TGF-beta regulates aromatase gene transcription, several reporter constructs containing different lengths of the placental specific promoter of the human aromatase gene were generated. JEG-3 cells were transiently transfected with a promoter construct and treated with or without TGF-beta1. The promoter activity was measured by luciferase assays. To examine the downstream signaling molecule mediating the effect of TGF-beta on aromatase transcription, cells were transiently transfected with dominant negative mutants of TGF-beta type II (TbetaRII) and type I receptor (ALK5) receptors before TGF-beta treatment. Smad2 activation was assessed by measuring phophorylated Smad2 protein levels in cytosolic and nuclear fractions. Smad2 expression was silenced using a siRNA expression construct. Finally, aromatase mRNA half-life was determined by treating cells with actinomycin D together with TGF-beta1 and measuring aromatase mRNA levels at various time points after treatment. Results and Discussion: TGF-beta1 inhibited the aromatase promoter activity in a time- and dosedependent manner. Deletion analysis suggests that the TGF-β1 response element resides between -422 and -117 nucleotides upstream from the transcription start site where a Smad binding element was found. The inhibitory effect of TGF-beta1 was blocked by dominant negative mutants of TbetaRII and ALK5. TGFbeta1 treatment induced Smad2 phosphorylation and translocation into the nucleus. On the other hand, kn
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