Increased acid ceramidase expression depends on upregulation of androgen-dependent deubiquitinases, USP2, in a human prostate cancer cell line, LNCaP
Increased acid ceramidase expression depends on upregulation of androgen-dependent deubiquitinases, USP2, in a human prostate cancer cell line, LNCaP
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DOI：
10.1093/jb/mvv039
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发表时间：
2015-10-01
影响因子：
2.7
通讯作者：
Murate, Takashi
中科院分区：
文献类型：
--
作者：
Mizutani, Naoki;Inoue, Minami;Murate, Takashi
酸性神经酰胺酶 (ACDase) 将神经酰胺代谢为鞘氨醇，从而产生 1-磷酸鞘氨醇。据报道，ACDase 在前列腺癌中表达上调。然而，其调控机制仍不清楚。 LNCaP（雄激素敏感的前列腺癌细胞系）而非 PC3 和 DU-145（雄激素无反应细胞系）表现出最高的 ACDase 蛋白。在三个细胞系中，ASAH1 mRNA 水平与 ACDase 蛋白表达不相关，并且 5'-启动子活性不表现出雄激素依赖性，表明 LNCaP 细胞中 ACDase 的转录后调节。根据这些结果，进一步分析了 LNCaP。 Casodex、雄激素受体拮抗剂和炭剥离 FCS (CS-FCS) 降低了 ACDase 蛋白和酶活性，而 CS-FCS 培养物中的二氢睾酮则增加了 ACDase 蛋白和酶活性。 MG132 是一种蛋白酶体抑制剂，在 CS-FCS 中培养时可阻止 ACDase 蛋白的减少，表明泛素/蛋白酶体系统参与其中。据报道，USP2是一种去泛素酶，在LNCaP细胞中发挥着重要作用。 USP2 siRNA 降低了 LNCaP 细胞的 ACDase 蛋白，而 USP2 过表达则增加了 LNCaP 细胞的 ACDase 蛋白。然而，SKP2（一种已知在前列腺癌中具有活性的泛素 E3 连接酶）并不影响 LNCaP 细胞中雄激素依赖性 ACDase 的表达。因此，雄激素敏感的 LNCaP 细胞中雄激素对 ACDase 的调节主要是由于雄激素刺激的 USP2 表达延长了其蛋白质半衰期。
Acid ceramidase (ACDase) metabolizes ceramide to sphingosine, leading to sphingosine 1-phosphate production. Reportedly, ACDase has been upregulated in prostate cancer. However, its regulatory mechanism remains unclear. LNCaP (androgen-sensitive prostate cancer cell line) but not PC3 and DU-145, (androgen-unresponsive cell lines) exhibited the highest ACDase protein. Among three cell lines, ASAH1 mRNA level was not correlated with ACDase protein expression, and the 5'-promoter activity did not show androgen dependency, suggesting the post-transcriptional regulation of ACDase in LNCaP cells. Based on these results, LNCaP was analysed further. Casodex, androgen receptor antagonist, and charcoal-stripped FCS (CS-FCS) decreased ACDase protein and activity, whereas dihydrotestosterone in CS-FCS culture increased ACDase protein and enzyme activity. MG132, a proteasome inhibitor, prevented the decrease of ACDase protein when cultured in CS-FCS, suggesting the involvement of ubiquitin/proteasome system. Reportedly, USP2, a deubiquitinase, plays an important role in LNCaP cells. USP2 siRNA decreased ACDase protein, whereas USP2 overexpression increased ACDase protein of LNCaP cells. However, SKP2, an ubiquitin E3 ligase known to be active in prostate cancer, did not affect androgen-dependent ACDase expression in LNCaP cells. Thus, ACDase regulation by androgen in androgen-sensitive LNCaP cells is mainly due to its prolonged protein half-life by androgen-stimulated USP2 expression.