Expression of RFG/ELE1alpha/ARA70 in normal and malignant prostatic epithelial cell cultures and lines: regulation by methylation and sex steroids.
RET fused gene (RFG)/ELE1alpha/androgen receptor-associated protein 70(ARA70) was first found to be involved in the activation of the RET proto-oncogene in thyroid neoplasm and has recently been shown to be a ligand-dependent transcriptional coregulator for androgen receptor (AR). The functionality of RFG/ELE1alpha/ARA70 remains controversial, and little is known about factors regulating its expression in the prostate. Of significant interest is whether this molecule is involved in prostate carcinogenesis. Using reverse transcriptase-polymerase chain reaction semiquantitation, we compared RFG/ELE1alpha/ARA70 mRNA levels in four prostate cancer cell lines (LNCaP, TSU-Pr1, DU-145, and PC-3) with those found in primary cultures of normal prostatic epithelial cells (PrECs). In addition, we examined the effects of androgen and antiandrogen, estrogen and antiestrogen, and a demethylating agent on RFG/ELE1alpha/ARA70 mRNA expression levels in AR- and AR+ PC-3 cells. Reduced levels of RFG/ELE1alpha/ARA70 message were observed in all four prostate cancer cell lines when compared with normal PrECs in primary cultures. RFG/ELE1alpha/ARA70 mRNA levels in PC-3 cells, which express both estrogen receptor subtypes, were upregulated by 17beta-estradiol and inhibited by the antiestrogen ICI-182780. In PC-3(AR+) cells, which were genetically engineered to express AR, exposure to androgen upregulated RFG/ELE1alpha/ARA70 mRNA expression, whereas treatment with 4-hydroxyflutamide lowered expression of this transcript. Furthermore, treatment of DU-145 cells, which did not express RFG/ELE1alpha/ARA70 transcripts, with a demethylating agent reactivated transcription of this gene. Polymerase chain reaction analyses of monochromosomal human-rodent hybrid panels localized a putative RFG/ELE1alpha/ARA70 isoform on human chromosome 5q31.1-31.2. In summary, we identified sex hormones and DNA hypermethylation as regulators of RFG/ELE1alpha/ARA70 expression in prostate cancer cells. In addition, we found reduced levels of RFG/ELE1alpha/ARA70 expression in prostate cancer cell lines when compared with expression levels in normal PrECs in culture. These findings suggest that RFG/ELE1alpha/ARA70 may be involved prostate carcinogenesis and that it may serve as a key mediator of estrogen-androgen synergism.
Division of Urology, Department of Surgery, University of Massachusetts Medical School, Worcester, Massachusetts 01655, USA.
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Tekur
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Lau
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Long
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Burnstein
S M
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Ho
D000728:Androgens / Q000502:physiology*; D001483:Base Sequence; D002874:Chromosome Mapping; D002895:Chromosomes, Human, Pair 5; D019175:DNA Methylation; D017931:DNA Primers; D004847:Epithelial Cells / Q000378:metabolism; D004967:Estrogens / Q000502:physiology*; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D006801:Humans; D008297:Male; D011467:Prostate / Q000166:cytology / Q000378:metabolism*; D011471:Prostatic Neoplasms / Q000235:genetics* / Q000473:pathology; D000090063:Proto-Oncogene Mas; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D000728:Androgens; D017931:DNA Primers; D004967:Estrogens; C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D011993:Recombinant Fusion Proteins