Sox7 通过 PSMA 增强子负调控前列腺特异性膜抗原 (PSMA) 的表达。
Sox7 negatively regulates prostate-specific membrane antigen (PSMA) expression through PSMA-enhancer.
机构信息
Winship Cancer Institute, Department of Hematology and Medical Oncology, Emory University School of Medicine, Atlanta, Georgia.
Department of Gastrointestinal Surgery, The Second Affiliated Hospital Of Soochow University, Suzhou, Jiangsu Province, P.R. China.
出版信息
Prostate. 2019 Mar;79(4):370-378. doi: 10.1002/pros.23743. Epub 2018 Nov 28.
BACKGROUND
PSMA expression in the prostate epithelium is controlled by a cis-element, PSMA enhancer (PSME). PSME contains multiple binding sites for Sox proteins, and in this study, we identified Sox7 protein as a negative regulator of PSMA expression through its interaction with PSME.
METHODS
The statistical correlation between Sox7 and PSMA mRNA expression was evaluated using five prostate cancer studies from cBioportal. In vitro and in vivo interaction between Sox7 and PSME was evaluated by chromatin immunoprecipitation (ChIP), electrophoretic mobility shift assay (EMSA), and luciferase reporter assay. Synthetic oligonucleotides were generated to define the sites in PSME that interact with Sox7 protein. Sox7 mutants were generated to identify the region of this protein required to regulate PSMA expression. Sox7 was also stably expressed in LNCaP/C4-2 and 22Rv1 cells to validate the regulation of PSMA expression by Sox7 in vivo.
RESULTS
Sox7 mRNA expression negatively correlated with PSMA/FOLH1 and PSMAL/FOLH1B mRNA expression in Broad/Cornell, TCGA and MSKCC studies, but not in two studies containing only metastatic prostate tumors. PC-3 cells mostly expressed the 48.5 KDa isoform 2 of Sox7, and the depletion of this isoform did not restore PSMA expression. Ectopic expression of canonical, wild-type Sox7 in C4-2 and 22Rv1 cells suppressed PSMA protein expression. ChIP assay revealed that canonical Sox7 protein preferentially interacts with PSME in vivo, and EMSA identified the SOX box sites #2 and #4 in PSME as required for its interaction. Sox7 was capable of directly binding to PSME and suppressed PSME-mediated transcription. The NLS regions of Sox7, but not its β-catenin interacting motif, are essential for this suppressing activity. Furthermore, restoration of wild-type Sox7 expression but not Sox7-NLS mutant in Sox7-null prostate cancer cell lines suppressed PSMA expression.
CONCLUSIONS
The inactivation of canonical Sox7 is responsible for the upregulated expression of PSMA in non-metastatic prostate cancer.
背景
前列腺上皮细胞中的 PSMA 表达受顺式元件 PSMA 增强子（PSME）控制。PSME 包含多个 Sox 蛋白结合位点，在本研究中，我们通过与 PSME 的相互作用鉴定出 Sox7 蛋白是 PSMA 表达的负调控因子。
方法
使用 cBioportal 中的五个前列腺癌研究评估 Sox7 和 PSMA mRNA 表达之间的统计相关性。通过染色质免疫沉淀（ChIP）、电泳迁移率变动分析（EMSA）和荧光素酶报告基因分析评估 Sox7 和 PSME 之间的体外和体内相互作用。生成合成寡核苷酸以定义与 Sox7 蛋白相互作用的 PSME 中的位点。生成 Sox7 突变体以鉴定该蛋白调节 PSMA 表达所需的区域。还在 LNCaP/C4-2 和 22Rv1 细胞中稳定表达 Sox7，以体内验证 Sox7 对 PSMA 表达的调节作用。
结果
Sox7 mRNA 表达与 Broad/Cornell、TCGA 和 MSKCC 研究中的 PSMA/FOLH1 和 PSMAL/FOLH1B mRNA 表达呈负相关，但在仅包含转移性前列腺肿瘤的两项研究中则无相关性。PC-3 细胞主要表达 Sox7 的 48.5 kDa 同工型 2，而该同工型的耗竭并不能恢复 PSMA 表达。C4-2 和 22Rv1 细胞中经典野生型 Sox7 的异位表达抑制了 PSMA 蛋白表达。ChIP 分析显示，经典 Sox7 蛋白在体内优先与 PSME 相互作用，EMSA 鉴定出 PSME 中的 SOX 盒位点 #2 和 #4 是其相互作用所必需的。Sox7 能够直接与 PSME 结合并抑制 PSME 介导的转录。Sox7 的核定位信号（NLS）区域，但不是其 β-连环蛋白相互作用基序，对于这种抑制活性是必需的。此外，在 Sox7 缺失的前列腺癌细胞系中恢复野生型 Sox7 表达而不是 Sox7-NLS 突变体抑制了 PSMA 表达。
结论
经典 Sox7 的失活导致非转移性前列腺癌中 PSMA 的上调表达。
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