Department of Zoology and Physiology, University of Wyoming, Laramie, Wyoming, United States of America.
PLoS One. 2012;7(7):e41242. doi: 10.1371/journal.pone.0041242. Epub 2012 Jul 24.
The peptidylarginine deiminase (PAD) family of enzymes post-translationally convert positively charged arginine residues in substrate proteins to the neutral, non-standard residue citrulline. PAD family members 1, 2, 3, and 6 have previously been localized to the cell cytoplasm and, thus, their potential to regulate gene activity has not been described. We recently demonstrated that PAD2 is expressed in the canine mammary gland epithelium and that levels of histone citrullination in this tissue correlate with PAD2 expression. Given these observations, we decided to test whether PAD2 might localize to the nuclear compartment of the human mammary epithelium and regulate gene activity in these cells. Here we show, for the first time, that PAD2 is specifically expressed in human mammary gland epithelial cells and that a portion of PAD2 associates with chromatin in MCF-7 breast cancer cells. We investigated a potential nuclear function for PAD2 by microarray, qPCR, and chromatin immunoprecipitation analysis. Results show that the expression of a unique subset of genes is disregulated following depletion of PAD2 from MCF-7 cells. Further, ChIP analysis of two of the most highly up- and down-regulated genes (PTN and MAGEA12, respectively) found that PAD2 binds directly to these gene promoters and that the likely mechanism by which PAD2 regulates expression of these genes is via citrullination of arginine residues 2-8-17 on histone H3 tails. Thus, our findings define a novel role for PAD2 in gene expression in human mammary epithelial cells.
肽基精氨酸脱亚氨酶（PAD）家族的酶在翻译后将底物蛋白中的正电荷精氨酸残基转化为中性、非标准残基瓜氨酸。PAD 家族成员 1、2、3 和 6 以前定位于细胞质中，因此，它们调节基因活性的潜力尚未被描述。我们最近证明 PAD2 在犬乳腺上皮细胞中表达，并且该组织中组蛋白瓜氨酸化水平与 PAD2 表达相关。鉴于这些观察结果，我们决定测试 PAD2 是否可能定位于人乳腺上皮细胞核区室并调节这些细胞中的基因活性。在这里，我们首次表明 PAD2 特异性表达于人乳腺上皮细胞，并且 PAD2 的一部分与 MCF-7 乳腺癌细胞中的染色质相关联。我们通过微阵列、qPCR 和染色质免疫沉淀分析研究了 PAD2 的潜在核功能。结果表明，在 MCF-7 细胞中耗尽 PAD2 后，一组独特的基因表达失调。此外，对两个上调和下调最显著的基因（PTN 和 MAGEA12）的 ChIP 分析发现，PAD2 直接结合到这些基因启动子上，并且 PAD2 调节这些基因表达的可能机制是通过组蛋白 H3 尾部第 2-8-17 位精氨酸残基的瓜氨酸化。因此，我们的研究结果定义了 PAD2 在人乳腺上皮细胞基因表达中的新作用。
