Reactive oxygen species (ROS) function as cellular switches for signaling cascade involved in cell growth, cell death, mitogenesis, angiogenesis and carcinogenesis. ROS are produced as a byproduct of oxidative phosphorylation (OXPHOS) in the mitochondria. It is estimated that 2-4% of the oxygen consumed during OXPHOS is converted to ROS. Besides mitochondria, NADPH-oxidase 1 (Nox1) also generates a significant amount of ROS in the cell. In this paper, we tested the hypothesis that mitochondria control Nox 1 redox signaling and the loss of control of this signaling contribute to tumorigenesis. We analyzed Nox1 expression in a mitochondrial gene knockout (ρ 0 ) cell line and in the isogenic cybrid cell line in which mitochondrial genes were restored by transfer of wild type mitochondria into ρ 0 cells. Our study revealed, for the first time, that the inactivation of mitochondrial genes leads to down-regulation of Nox1 and that the transfer of wild type mitochondrial genes restored the Nox1 expression to a level comparable to that in the parental cell line. Consistent with Nox1 down-regulation, we found that ρ 0 cells contained low levels of superoxide anion and that superoxide levels reversed to parental levels in cybrid cells when Nox1 expression was restored by transfer of wild type mitochondria. Increasing mitochondrial superoxide levels also increased the expression of Nox1 in parental cells. Confocal microscopy studies revealed that Nox1 localizes in the mitochondria. Nox1 was highly expressed in breast (86%) and ovarian (71%) tumors and that its expression positively correlated with expression of cytochrome C oxidase encoded by mtDNA. Our study, described in this paper demonstrates the existence of cross talk between the mitochondria and NADPH oxidase. Furthermore, our studies suggest that mitochondria control Nox1 redox signaling and the loss of control of this signaling contributes to breast and ovarian tumorigenesis.

Estrogen receptor-␣ (ER␣) promotes proliferation of breast cancer cells, whereas tumor suppressor protein p53 impedes proliferation of cells with genomic damage. Whether there is a direct link between these two antagonistic pathways has remained unclear. Here we report that ER␣ binds directly to p53 and represses its function. The activation function-2 (AF-2) domain of ER␣ and the C-terminal regulatory domain of p53 are necessary for the interaction. Knocking down p53 and ER␣ by small interfering RNA elicits opposite effects on p53-target gene expression and cell cycle progression. Remarkably, ionizing radiation that causes genomic damage disrupts the interaction between ER␣ and p53. Ionizing radiation together with ER␣ knock down results in an additive effect on transcription of endogenous p53-target gene p21 (CDKN1) in human breast cancer cells. Our findings reveal a novel mechanism for regulating p53 and suggest that suppressing p53 function is an important component in the proproliferative role of ER␣.As a tumor suppressor, p53 plays a central role in cellular processes such as cell cycle arrest, apoptosis, senescence, and differentiation (1, 2). Although these functions of p53 are essential to prevent cells from becoming cancerous, left uncontrolled, they can lead to consequences deleterious to normal cells. Mutations in the p53 gene or aberrations in the mechanisms to balance p53 function pave the way to tumorigenesis (3). p53 elicits its biological functions mainly by functioning as a transcriptional regulator of various cellular genes with p53-response elements. On the other hand, estrogen receptor-␣ (ER␣) 5 regulates growth and development of various tissues and promotes proliferation of breast cancer cells (4 -8). ER␣ is a transcriptional regulator that is recruited to the promoter regions of target genes directly through binding to estrogen response elements (EREs) or indirectly through other DNA-binding factors, such as AP1 and Sp1 (7, 9). The opposing functions of p53 and ER␣, while stringently controlled in normal cells, are likely disrupted in cancer cells. Various observations have alluded to the potential for a cross-talk between p53 and ER␣ signaling pathways. For example, in murine models, early exposure to 17␤-estradiol (E 2 ) and progesterone to mimic pregnancy induced nuclear p53 enabling resistance to carcinogenesis by blocking proliferation of apparently ER␣-positive cells (10). In breast cancer cells, increased expression of ER␣ led to elevated levels of p53 and MDM2, an inhibitor of p53 function (11), whereas overexpression of MDM2 enhanced the function of ER␣ (12). However, whether there is a direct link between the p53 and ER␣ pathways has remained unclear. To address this important issue, we investigated whether ER␣ directly interacts with p53 and affects its function. EXPERIMENTAL PROCEDURESCell Culture and Irradiation-MCF7 cells and Saos2 cells were maintained in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum (FBS) (Invitrogen) or 10% dextran-charcoal-t...

Current standard-of-care (SOC) therapy for breast cancer includes targeted therapies such as endocrine therapy for estrogen receptor-alpha (ERα) positive; anti-HER2 monoclonal antibodies for human epidermal growth factor receptor-2 (HER2)-enriched; and general chemotherapy for triple negative breast cancer (TNBC) subtypes. These therapies frequently fail due to acquired or inherent resistance. Altered metabolism has been recognized as one of the major mechanisms underlying therapeutic resistance. There are several cues that dictate metabolic reprogramming that also account for the tumors’ metabolic plasticity. For metabolic therapy to be efficacious there is a need to understand the metabolic underpinnings of the different subtypes of breast cancer as well as the role the SOC treatments play in targeting the metabolic phenotype. Understanding the mechanism will allow us to identify potential therapeutic vulnerabilities. There are some very interesting questions being tackled by researchers today as they pertain to altered metabolism in breast cancer. What are the metabolic differences between the different subtypes of breast cancer? Do cancer cells have a metabolic pathway preference based on the site and stage of metastasis? How do the cell-intrinsic and -extrinsic cues dictate the metabolic phenotype? How do the nucleus and mitochondria coordinately regulate metabolism? How does sensitivity or resistance to SOC affect metabolic reprogramming and vice-versa? This review addresses these issues along with the latest updates in the field of breast cancer metabolism.

Estrogen receptor A (ERA) and tumor suppressor protein p53 exert opposing effects on cellular proliferation. As a transcriptional regulator, p53 is capable of activating or repressing various target genes. We have previously reported that ERA binds directly to p53, leading to down-regulation of transcriptional activation by p53. In addition to transcriptional activation, transcriptional repression of a subset of target genes by p53 plays important roles in diverse biological processes, such as apoptosis. Here, we report that ERA inhibits p53-mediated transcriptional repression. Chromatin immunoprecipitation assays reveal that ERA interacts in vivo with p53 bound to promoters of Survivin and multidrug resistance gene 1, both targets for transcriptional repression by p53. ERA binding to p53 leads to inhibition of p53-mediated transcriptional regulation of these genes in human cancer cells. Transcriptional derepression of Survivin by ERA is dependent on the p53-binding site on the Survivin promoter, consistent with our observation that p53 is necessary for ERA to access the promoters. Importantly, mutagenic conversion of this site to an activation element enabled ERA to repress p53-mediated transcriptional activation. Further, RNA interference-mediated knockdown of ERA resulted in reduced Survivin expression and enhanced the propensity of MCF-7 cells to undergo apoptosis in response to staurosporine treatment, an effect that was blocked by exogenous expression of Survivin. These results unravel a novel mechanism by which ERA opposes p53-mediated apoptosis in breast cancer cells. The findings could have translational implications in developing new therapeutic and prevention strategies against breast cancer. [Cancer Res 2007;67(16):7746-55]

Estrogen receptor α (ERα) plays an important role in the onset and progression of breast cancer, whereas p53 functions as a major tumor suppressor. We previously reported that ERα binds to p53, resulting in inhibition of transcriptional regulation by p53. Here, we report on the molecular mechanisms by which ERα suppresses p53's transactivation function. Sequential ChIP assays demonstrated that ERα represses p53-mediated transcriptional activation in human breast cancer cells by recruiting nuclear receptor corepressors (NCoR and SMRT) and histone deacetylase 1 (HDAC1). RNAi-mediated down-regulation of NCoR resulted in increased endogenous expression of the cyclin-dependent kinase (CDK)-inhibitor p21 Waf1/Cip1 (CDKN1A) gene, a prototypic transcriptional target of p53. While 17β-estradiol (E2) enhanced ERα binding to p53 and inhibited p21 transcription, antiestrogens decreased ERα recruitment and induced transcription. The effects of estrogen and antiestrogens on p21 transcription were diametrically opposite to their known effects on the conventional ERE-containing ERα target gene, pS2/TFF1. These results suggest that ERα uses dual strategies to promote abnormal cellular proliferation: enhancing the transcription of ERE-containing proproliferative genes and repressing the transcription of p53-responsive antiproliferative genes. Importantly, ERα binds to p53 and inhibits transcriptional activation by p53 in stem/progenitor cell-containing murine mammospheres, suggesting a potential role for the ER-p53 interaction in mammary tissue homeostasis and cancer formation. Furthermore, retrospective studies analyzing response to tamoxifen therapy in a subset of patients with ER-positive breast cancer expressing either wild-type or mutant p53 suggest that the presence of wild-type p53 is an important determinant of positive therapeutic response.nuclear receptor corepressor | mammary epithelial cells | mammospheres | tumor suppressor protein | tamoxifen therapy