Estrogen receptor (ER) a and ERR are present in mitochondria but little is known about their contribution to the overall constellation of cellular responses to 17 beta-estradiol, which inhibits the early stages of apoptosis through membrane and mitochondrial, but not nuclear, ER signaling pathways. Whereas membrane ER signaling leading to inhibition of apoptosis involves the activation of protein kinase pathways, mitochondrial ER-mediated inhibition involves the direct and/or indirect activation of manganese superoxide dismutase by the mitochondrial ER.

Our previous studies have shown that 17beta estradiol (E2) enhances the transcript levels of mitochondrial DNA (mtDNA)-encoded genes and mitochondrial respiratory chain (MRC) activity via estrogen receptors (ER). Others have reported the presence of putative estrogen responsive elements (ERE) in human mtDNA (mtEREs) and detection of ERs in mitochondria of rat uterine and ovary cells. Recently, we demonstrated the E2-enhanced mitochondrial localization of ERalpha and ERbeta, and E2-induced mtDNA transcript levels in MCF-7 cells. In this study, we applied electrophoresis mobility shift assays (EMSAs) and surface plasmon resonance (SPR) to determine if mitochondrial extracts, recombinant human ERalpha (rhERalpha), and rhERbeta interact with mtEREs. Using EMSAs, we observed that ER-containing mitochondrial extracts bound to mtEREs and the binding was enhanced by E2, whereas the binding of mitochondrial proteins from ERbeta-deficient cells was almost undetectable. Both rhERalpha and rhERbeta bound to the mtEREs and their binding was altered by their respective antibodies. However, the ERa antibodies did not alter the binding of MCF-7 cell mitochondrial extracts to mtEREs whereas the binding MCF-7 and MDA-MB-231 cell mitochondrial extracts to mtEREs was reduced by ERbeta antibody. These results suggest that the mtERE-bound mitochondrial protein is ERbeta. Using SPR, we observed the binding of both ERs to mtEREs and that the binding was increased by E2. These results indicate that the mitochondrial ERs can interact with mtEREs and suggest that they may be directly involved in E2 induction of mtDNA transcription. (C) 2004 Wiley-Liss, Inc.

We observed previously that estrogen treatment increased the transcript levels of several mitochondrial DNA (mtDNA)-encoded genes for mitochondrial respiratory chain (MRC) proteins and MRC activity in rat hepatocytes and human Hep G2 cells. Others have reported detection of estrogen receptors (ER), ERα and ERβ, in mitochondria of rabbit ovarian and uterine tissue. In this study, we have extended these observations. Using cellular fractionation and Western blot with ERα- and ERβ-specific antibodies, we observed that ERα and ERβ are present in mitochondria of human MCF7 cells and that the mitochondrial ERα and ERβ account for 10 and 18%, respectively, of total cellular ERα and ERβ in 17β-estradiol (E2)-treated MCF7 cells. We also found that E2 significantly enhanced the amounts of mitochondrial ERα and ERβ in a time- and concentration-dependent manner and that these effects are accompanied by a significant increase in the transcript levels of mtDNA-encoded genes, i.e., cytochrome c oxidase subunits I and II. Moreover, we demonstrated that these E2-mediated effects were inhibited by the pure ER antagonist, ICI-182780, indicating the involvement of ERs. Using immunohistochemistry with confocal microscopy and immunogold electron microscopy, we demonstrated that ERα and ERβ are located within the MCF7 cell mitochondrial matrix. Computer analysis identified a putative internal mitochondrial targeting peptide signal within human ERβ, suggesting an inherent potential for ERβ to enter mitochondria. These findings confirm the observations of others and provide additional support for this novel localization of the ERs and for a potentially important role of the ER in the regulation of mtDNA transcription.

Chen, Jin Q., Michael Delannoy, Carol Cooke, and James D.Yager. Mitochondrial localization of ER and ER in human MCF7cells. Am J Physiol Endocrinol Metab 286: E1011–E1022, 2004. Firstpublished January 21, 2004; 10.1152/ajpendo.00508.2003.—We ob-served previously that estrogen treatment increased the transcriptlevels of several mitochondrial DNA (mtDNA)-encoded genes formitochondrial respiratory chain (MRC) proteins and MRC activity inrat hepatocytes and human Hep G2 cells. Others have reporteddetection of estrogen receptors (ER), ER and ER , in mitochondriaof rabbit ovarian and uterine tissue. In this study, we have extendedthese observations. Using cellular fractionation and Western blot withER - and ER -specific antibodies, we observed that ER and ERare present in mitochondria of human MCF7 cells and that themitochondrial ER and ER account for 10 and 18%, respectively, oftotal cellular ER and ER in 17 -estradiol (E

We observed previously that estrogen treatment increased the transcript levels of several mitochondrial DNA (mtDNA)-encoded genes for mitochondrial respiratory chain (MRC) proteins and MRC activity in rat hepatocytes and human Hep G2 cells. Others have reported detection of estrogen receptors (ER), ERalpha and ERbeta, in mitochondria of rabbit ovarian and uterine tissue. In this study, we have extended these observations. Using cellular fractionation and Western blot with ERalpha- and ERbeta-specific antibodies, we observed that ERalpha and ERbeta are present in mitochondria of human MCF7 cells and that the mitochondrial ERalpha and ERbeta account for 10 and 18%, respectively, of total cellular ERalpha and ERbeta in 17beta-estradiol (E(2))-treated MCF7 cells. We also found that E(2) significantly enhanced the amounts of mitochondrial ERalpha and ERbeta in a time- and concentration-dependent manner and that these effects are accompanied by a significant increase in the transcript levels of mtDNA-encoded genes, i.e., cytochrome c oxidase subunits I and II. Moreover, we demonstrated that these E(2)-mediated effects were inhibited by the pure ER antagonist, ICI-182780, indicating the involvement of ERs. Using immunohistochemistry with confocal microscopy and immunogold electron microscopy, we demonstrated that ERalpha and ERbeta are located within the MCF7 cell mitochondrial matrix. Computer analysis identified a putative internal mitochondrial targeting peptide signal within human ERbeta, suggesting an inherent potential for ERbeta to enter mitochondria. These findings confirm the observations of others and provide additional support for this novel localization of the ERs and for a potentially important role of the ER in the regulation of mtDNA transcription.