Acid Ceramidase Promotes Nuclear Export of PTEN through Sphingosine 1-Phosphate Mediated Akt Signaling

The tumor suppressor PTEN is now understood to regulate cellular processes at the cytoplasmic membrane, where it classically regulates PI3K signaling, as well as in the nucleus where multiple roles in controlling cell cycle and genome stability have been elucidated. Mechanisms that dictate nuclear import and, less extensively, nuclear export of PTEN have been described, however the relevance of these processes in disease states, particularly cancer, remain largely unknown. We investigated the impact of acid ceramidase on the nuclear-cytoplasmic trafficking of PTEN. Immunohistochemical analysis of a human prostate tissue microarray revealed that nuclear PTEN was lost in patients whose tumors had elevated acid ceramidase. We found that acid ceramidase promotes a reduction in nuclear PTEN that is dependent upon sphingosine 1-phosphate-mediated activation of Akt. We were further able to show that sphingosine 1-phosphate promotes formation of a complex between Crm1 and PTEN, and that leptomycin B prevents acid ceramidase and sphingosine 1-phosphate mediated loss of nuclear PTEN, suggesting an active exportin-mediated event. To investigate whether the tumor promoting aspects of acid ceramidase in prostate cancer depend upon its ability to export PTEN from the nucleus, we used enforced nuclear expression of PTEN to study docetaxel-induced apoptosis and cell killing, proliferation, and xenoengraftment. Interestingly, while acid ceramidase was able to protect cells expressing wild type PTEN from docetaxel, promote proliferation and xenoengraftment, acid ceramidase had no impact in cells expressing PTEN-NLS. These findings suggest that acid ceramidase, through sphingosine 1-phosphate, promotes nuclear export of PTEN as a means of promoting tumor formation, cell proliferation, and resistance to therapy.

目前学界已明确，抑癌蛋白PTEN（PTEN）可在细胞质膜上调控多种细胞进程，其经典功能为调控PI3K信号通路；同时在细胞核内也发挥重要作用，目前已阐明其在调控细胞周期与基因组稳定性方面存在多种功能。目前已有研究阐明了调控PTEN入核的分子机制，而关于其出核的机制报道相对较少；但上述转运过程在疾病状态尤其是癌症中的相关生物学意义，目前仍基本不明。本研究探讨了酸性神经酰胺酶（acid ceramidase）对PTEN核质穿梭的影响。对人类前列腺组织微阵列开展的免疫组化分析结果显示，肿瘤组织中酸性神经酰胺酶表达升高的患者，其细胞核内PTEN表达缺失。本研究发现，酸性神经酰胺酶可通过依赖鞘氨醇1-磷酸（sphingosine 1-phosphate）介导的蛋白激酶B（Akt）激活通路，促进细胞核内PTEN水平降低。本研究进一步证实，鞘氨醇1-磷酸可促进Crm1（染色体区域维持蛋白1）与PTEN形成复合物；而细霉素B（leptomycin B）可阻断酸性神经酰胺酶与鞘氨醇1-磷酸介导的细胞核内PTEN水平降低，这表明该过程为依赖输出蛋白的主动转运事件。为探究酸性神经酰胺酶在前列腺癌中的促瘤作用是否依赖其介导PTEN出核的能力，本研究通过使PTEN定向定位于细胞核的强制表达策略，研究了多西他赛（docetaxel）诱导的细胞凋亡、细胞杀伤效应、细胞增殖以及异种移植（xenoengraftment）情况。有趣的是，尽管酸性神经酰胺酶可保护表达野生型PTEN的细胞免受多西他赛的杀伤，同时促进细胞增殖与异种移植，但在表达PTEN-NLS（PTEN-NLS）的细胞中，酸性神经酰胺酶并未产生任何影响。上述研究结果表明，酸性神经酰胺酶可通过鞘氨醇1-磷酸介导PTEN的核输出，以此促进肿瘤发生、细胞增殖以及治疗耐药性的产生。