Matsuzaki Youichirou, Koyama Makoto, Hitomi Toshiaki, Takaoka Yuuki, Kawanaka Mayumi, Sakai Toshiyuki
Department of Molecular-Targeting Cancer Prevention, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kawaramachi-Hirokoji, Kyoto 602-8566, Japan.
Int J Oncol. 2005 Aug;27(2):497-503.
15-deoxy-Delta12, 14-prostaglandin J2 (15-d-PGJ2) inhibits cellular proliferation primarily in the G1 phase of the cell cycle. However, the molecular mechanism responsible for this effect has not been sufficiently elucidated. Here, we show that the treatment of human immortalized keratinocyte HaCaT cells with 15-d-PGJ2 arrests the cell cycle at the G1 phase. We also show that inhibition of the proliferation of HaCaT cells by 15-d-PGJ2 correlates with induction of the expression of p15INK4b protein, a cyclin-dependent kinase inhibitor. 15-d-PGJ2 also induces p15INK4b mRNA and its promoter activity, suggesting that 15-d-PGJ2 transcriptionally activates p15INK4b gene expression. Deletion and mutation analyses indicated that one of the elements responsible for 15-d-PGJ2-mediated induction is located between nucleotides -385 and -373 upstream of the initiation codon. An electrophoretic mobility shift assay revealed the specific binding of an unknown protein to this element, and that the binding is augmented by the treatment of 15-d-PGJ2. Although 15-d-PGJ2 is a natural ligand of the peroxisome proliferator-activated receptor gamma (PPARgamma), treatment with ciglitazone, a synthetic PPARgamma ligand, had no effect on p15INK4b gene expression. Collectively, these results indicate that 15-d-PGJ2 activates p15INK4b gene expression through a novel 15-d-PGJ2-responsive element in the p15INK4b promoter in a PPARgamma-independent manner.
15-脱氧-Δ12,14-前列腺素J2（15-d-PGJ2）主要在细胞周期的G1期抑制细胞增殖。然而，导致这种效应的分子机制尚未得到充分阐明。在此，我们表明用15-d-PGJ2处理人永生化角质形成细胞HaCaT细胞会使细胞周期停滞在G1期。我们还表明，15-d-PGJ2对HaCaT细胞增殖的抑制与细胞周期蛋白依赖性激酶抑制剂p15INK4b蛋白表达的诱导相关。15-d-PGJ2还诱导p15INK4b mRNA及其启动子活性，表明15-d-PGJ2转录激活p15INK4b基因表达。缺失和突变分析表明，负责15-d-PGJ2介导诱导的元件之一位于起始密码子上游-385至-373核苷酸之间。电泳迁移率变动分析揭示了一种未知蛋白质与该元件的特异性结合，并且这种结合通过15-d-PGJ2处理而增强。尽管15-d-PGJ2是过氧化物酶体增殖物激活受体γ（PPARγ）的天然配体，但用合成PPARγ配体吡格列酮处理对p15INK4b基因表达没有影响。总体而言，这些结果表明，15-d-PGJ2通过p15INK4b启动子中一个新的15-d-PGJ2反应元件以不依赖PPARγ的方式激活p15INK4b基因表达。
