Abstract
Evidence supporting an early origin of prostate cancer is growing. We demonstrated previously that brief exposure of neonatal rats to estradiol or bisphenol A elevated their risk of developing precancerous lesions in the prostate upon androgen-supported treatment with estradiol as adults. Epigenetic reprogramming may be a mechanism underlying this inductive event in early life, because we observed overexpression of phosphodiesterase 4D variant 4 (Pde4d4) through induction of hypomethylation of its promoter. This epigenetic mark was invisible in early life (postnatal d 10), becoming apparent only after sexual maturation. Here, we asked whether other estrogen-reprogrammable epigenetic marks have similar or different patterns in gene methylation changes throughout life. We found that hypomethylation of the promoter of nucleosome binding protein-1 (Nsbp1), unlike Pde4d4, is an early and permanent epigenetic mark of neonatal exposure to estradiol/bisphenol A that persists throughout life, unaffected by events during adulthood. In contrast, hippocalcin-like 1 (Hpcal1) is a highly plastic epigenetic mark whose hypermethylation depends on both type of early-life exposure and adult-life events. Four of the eight genes involved in DNA methylation/demethylation showed early and persistent overexpression that was not a function of DNA methylation at their promoters, including genes encoding de novo DNA methyltransferases (Dnmt3a/b) and methyl-CpG binding domain proteins (Mbd2/4) that have demethylating activities. Their lifelong aberrant expression implicates them in early-life reprogramming and prostate carcinogenesis during adulthood. We speculate that the distinctly different fate of early-life epigenetic marks during adulthood reflects the complex nature of lifelong editing of early-life epigenetic reprogramming.
Highlights
Evidence supporting an early origin of prostate cancer is growing
Apropos to early-life transcriptome remodeling through epigenetics in the prostate, we provided the first evidence that brief exposure of neonatal rats to 17-estradiol-3-benzoate (EB) or the environmental estrogen mimic bisphenol A (BPA) induced hypomethylation of the promoter of phosphodiesterase 4D variant 4 (Pde4d4) and its overexpression in their prostates during adulthood [23]
Real-time PCR As previously described [23, 24], total RNA was isolated from cells or tissues reverse transcribed to cDNA; and expression levels of nucleosome binding protein-1 (Nsbp1), hippocalcin-like 1 (Hpcal1), ribosomal protein L19 (Rpl19), Dnmt1, DNA methyltransferase-3a (Dnmt3a), Dnmt3b, methyl CpG binding protein 2 (Mecp2), Mbd1, methyl-CpG binding domain protein 2 (Mbd2), Mbd3, and Mbd4 determined by a SYBR Green-based real-time PCR method on the ABI PRISM 7500 FAST System (Applied Biosystems, Foster City, CA)
Summary
Animals and tissue preparation Animal housing and treatments were approved by the Animal. Real-time PCR As previously described [23, 24], total RNA was isolated from cells or tissues reverse transcribed to cDNA; and expression levels of Nsbp, Hpcal, ribosomal protein L19 (Rpl19), Dnmt, Dnmt3a, Dnmt3b, methyl CpG binding protein 2 (Mecp2), Mbd, Mbd, Mbd, and Mbd determined by a SYBR Green-based real-time PCR method on the ABI PRISM 7500 FAST System (Applied Biosystems, Foster City, CA). Primers were designed to amplify fragments encompassing the 5Ј-CGI of candidate genes: Nsbp, Hpcal, Dnmt3b, and Mbd from bisulfite-modified DNA (Table 1). To determine whether the differentially methylated genes can regulate cellular functions in rat prostatic epithelial cells, we used siRNA to knockdown gene expression in NbE-1 or AIT cells and studied the effects on cell viability. RNA was isolated for real-time PCR analysis of target-gene expression [23, 24].
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