Department of Breast Medical Oncology, Harbin Medical University Cancer Hospital, Harbin, Heilongjiang, P. R. China.
Department of Pathology, Harbin Medical University Cancer Hospital, Harbin, Heilongjiang, P. R. China.
Cancer Commun (Lond). 2023 May;43(5):582-612. doi: 10.1002/cac2.12417. Epub 2023 Apr 2.
Nuclear Yes1-associated transcriptional regulator (YAP1) promotes tumor progression. However, the function of cytoplasmic YAP1 in breast cancer cells and its impact on the survival of breast cancer patients remain unclear. Our research aimed to explore the biological function of cytoplasmic YAP1 in breast cancer cells and the possibility of cytoplasmic YAP1 as a predictive marker of breast cancer survival.
We constructed cell mutant models, including NLS-YAP1 (nuclear localized), YAP1 (incapable of binding to the TEA domain transcription factor family) and YAP1 (cytoplasmic localized), and used Cell Counting Kit-8 (CCK-8) assays, 5-ethynyl-2'-deoxyuridine (EdU) incorporation assays, and Western blotting (WB) analysis to detect cell proliferation and apoptosis. The specific mechanism of cytoplasmic YAP1-mediated endosomal sorting complexes required for transport III (ESCRT-III) assembly was studied by co-immunoprecipitation, immunofluorescence staining, and WB analysis. Epigallocatechin gallate (EGCG) was used to simulate YAP1 retention in the cytoplasm in in vitro and in vivo experiments to study the function of cytoplasmic YAP1. YAP1 binding to NEDD4-like E3 ubiquitin protein ligase (NEDD4L) was identified using mass spectrometry and was verified in vitro. Breast tissue microarrays were used to analyze the relationship between cytoplasmic YAP1 expression and the survival of breast cancer patients.
YAP1 was mainly expressed in the cytoplasm in breast cancer cells. Cytoplasmic YAP1 promoted autophagic death of breast cancer cells. Cytoplasmic YAP1 bound to the ESCRT-III complex subunits charged multivesicular body protein 2B (CHMP2B) and vacuolar protein sorting 4 homolog B (VPS4B), promoting assembly of CHMP2B-VPS4B and activating autophagosome formation. EGCG retained YAP1 in the cytoplasm, promoting the assembly of CHMP2B-VPS4B to promote autophagic death of breast cancer cells. YAP1 bound to NEDD4L, and NEDD4L mediated ubiquitination and degradation of YAP1. Breast tissue microarrays revealed that high levels of cytoplasmic YAP1 were beneficial to the survival of breast cancer patients.
Cytoplasmic YAP1 mediated autophagic death of breast cancer cells by promoting assembly of the ESCRT-III complex; furthermore, we established a new breast cancer survival prediction model based on cytoplasmic YAP1 expression.
核 Yes1 相关转录调节因子（YAP1）促进肿瘤进展。然而，细胞质 YAP1 在乳腺癌细胞中的功能及其对乳腺癌患者生存的影响尚不清楚。我们的研究旨在探讨细胞质 YAP1 在乳腺癌细胞中的生物学功能，以及细胞质 YAP1 作为乳腺癌生存预测标志物的可能性。
我们构建了细胞突变模型，包括核定位的 NLS-YAP1、不能与 TEA 结构域转录因子家族结合的 YAP1（YAP1）和细胞质定位的 YAP1，并用细胞计数试剂盒-8（CCK-8）检测试剂盒、5-乙炔基-2'-脱氧尿苷（EdU）掺入检测试剂盒和 Western blot（WB）分析检测细胞增殖和凋亡。通过共免疫沉淀、免疫荧光染色和 WB 分析研究了细胞质 YAP1 介导的内体分选复合物所需的运输 III（ESCRT-III）组装的具体机制。用表没食子儿茶素没食子酸酯（EGCG）在体外和体内实验中模拟 YAP1 在细胞质中的保留，以研究细胞质 YAP1 的功能。使用质谱法鉴定 YAP1 与 NEDD4 样 E3 泛素蛋白连接酶（NEDD4L）的结合，并在体外进行验证。使用乳腺癌组织微阵列分析细胞质 YAP1 表达与乳腺癌患者生存之间的关系。
YAP1 在乳腺癌细胞中主要表达在细胞质中。细胞质 YAP1 促进乳腺癌细胞的自噬性死亡。细胞质 YAP1 与 ESCRT-III 复合物亚基多泡体蛋白 2B（CHMP2B）和液泡蛋白分选 4 同源 B（VPS4B）结合，促进 CHMP2B-VPS4B 的组装，并激活自噬体形成。EGCG 将 YAP1 保留在细胞质中，促进 CHMP2B-VPS4B 的组装，从而促进乳腺癌细胞的自噬性死亡。YAP1 与 NEDD4L 结合，NEDD4L 介导 YAP1 的泛素化和降解。乳腺癌组织微阵列显示，高水平的细胞质 YAP1 有利于乳腺癌患者的生存。
细胞质 YAP1 通过促进 ESCRT-III 复合物的组装介导乳腺癌细胞的自噬性死亡；此外，我们基于细胞质 YAP1 表达建立了新的乳腺癌生存预测模型。
