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Interferons exert their biological function mainly through the activation of interferon-stimulated genes (ISGs). ISG12 (originally designated p27) belongs to a family of small, interferon a inducible genes of unknown function. We have determined the 5 0 end sequence of ISG12 cDNA from the human cell lines HeLa and AMA by RACE. Comparing this sequence to ISG12 sequences in the expressed sequence tag (EST) database revealed the presence of two alternative splice variants of ISG12 in human cells exhibiting the same open reading frame. We have sequenced the promoter region of the ISG12 gene and found ISRE, IRF1/IRF2, and STAT elements correlating to the interferon a inducibility of the gene. Subsequently, we have expressed human ISG12, a 12-kDa hydrophobic protein in the baculovirus expression system and with a C-terminal FLAG-tag in the human cell line 293. Recombinant ISG12 sediments in the nuclear envelope in both cell types. Finally, we have been able to demonstrate the prevalence of the ISG12 gene product in the nuclear envelope of HeLa cells treated with interferon a by immunocytochemical analyses. ISG12 is the first interferon induced protein found localizing to the nuclear envelope.Keywords: interferon; nuclear envelope; ISG12; baculovirus; FLAG.Interferons (IFNs) are a family of cytokines in vertebrates with a variety of biological functions, such as growth inhibition, antitumoral, immunomodulatory, antiviral, and antiparasitic actions. Human IFNs are grouped into type I and type II, and the biological actions of IFNs are mainly mediated through the gene products of more than 100 IFN stimulated genes (ISGs) (reviewed in [1,2]). The best characterized ISGs are PKR, the 2 0 -5 0 oligoadenylate synthetases (2-5OAS), and RNAse L (reviewed in [3 -5]). A group of small ISGs consisting of ISG12, 6-16, 9-27, 1-8, ISG15 and 6-26 still remains relatively uncharacterized and their functions are still basically unknown [6][7][8][9][10]. The 6-16 promoter has been well characterized and utilized in many studies [11,12], most prominently in the discovery of the JAK-STAT pathways [1,13].The cDNA of ISG12 (encoding a putative ISG protein of M r 12 000) was originally cloned as an oestrogen-induced gene in the human breast epithelial cell line, MCF7 and designated p27 [10]. Even though it was demonstrated that the level of expression of ISG12 mRNA is induced by oestrogen in MCF7 cells, and high expression levels of ISG12 mRNA are found in many breast carcinomas, ISG12 expression does not correlate with the presence of the oestrogen receptor neither in the cell lines tested nor in breast carcinomas [10]. Analysis of a number of breast tissue sections for expression of ISG12 mRNA by in situ hybridization revealed that: (a) no ISG12 transcripts were found in normal cells of the sections; (b) a high level of expression was found in cancer cells in most of the sections; and (c) a low level of expression was found in fibroblasts of some fibroadenomas [10]. In addition, we have previously demonstrated by quantitative RT-PCR that I...

IntroductionThe incidence of breast cancer has been increasing steadily over the past 60 years [1], today affecting one in eight women in the United States [2]. Despite tremendous efforts to understand the disease, less than 50% of all cases are of known etiology such as genetic inheritance, first-degree relatives with breast cancer, and age of menstruation and menopause [3].A large group of lipophilic organochlorines are found to persist in the environment and to biomagnify through the food chain. This group includes pesticides, polychlorinated biphenyls (PCBs) and polychlorinated dibenzo-dioxins/ polychlorinated dibenzo-furans [4][5][6][7][8]. Many of these compounds can interfere with a wide range of hormonal responses [9][10][11][12][13], including estrogen receptor (ER)-mediated responses. Due to their lipophilic nature, bp = base pair; DMEM = Dulbecco's modified Eagle's medium; E2 = 17β-estradiol; ER = estrogen receptor; PCB = polychlorinated biphenyl; PCR = polymerase chain reaction; POC = persistent organochlorine; TCDD = 2,3,7,8 tetrachlorodibenzo-p-dioxin.Available online http://breast-cancer-research.com/content/4/6/R12 Abstract Background: Environmental persistent organochlorines (POCs) biomagnify in the food chain, and the chemicals are suspected of being involved in a broad range of human malignancies. It is speculated that some POCs that can interfere with estrogen receptor-mediated responses are involved in the initiation and progression of human breast cancer. The tumor suppressor gene BRCA1 plays a role in cell-cycle control, in DNA repair, and in genomic stability, and it is often downregulated in sporadic mammary cancers. The aim of the present study was to elucidate whether POCs have the potential to alter the expression of BRCA1. Methods: Using human breast cancer cell lines MCF-7 and MDA-MB-231, the effect on BRCA1 expression of chemicals belonging to different classes of organochlorine chemicals (the pesticide toxaphene, 2,3,7,8-tetrachlorodibenzo-p-dioxin, and three polychlorinated biphenyls [PCB#138, PCB#153 and PCB#180]) was measured by a reporter gene construct carrying 267 bp of the BRCA1 promoter. A twofold concentration range was analyzed in MCF-7, and the results were supported by northern blot analysis of BRCA1 mRNA using the highest concentrations of the chemicals. Results: All three polychlorinated biphenyls and 2,3,7,8-tetrachlorodibenzo-p-dioxin reduced 17β-estradiol (E2)-induced expression as well as basal reporter gene expression in both cell lines, whereas northern blot analysis only revealed a downregulation of E2-induced BRCA1 mRNA expression in MCF-7 cells. Toxaphene, like E2, induced BRCA1 expression in MCF-7.Conclusion: The present study shows that some POCs have the capability to alter the expression of the tumor suppressor gene BRCA1 without affecting the cell-cycle control protein p21 Waf/Cip1 . Some POCs therefore have the potential to affect breast cancer risk.