PO.TB07.02 · 肿瘤生物学
PSMD2抑制破坏自噬周转并在ER+乳腺癌中形成自噬依赖性脆弱点
PSMD2 Suppression Disrupts Autophagy Turnover and Creates an Autophagy-Dependent Vulnerability in ER+ Breast Cancer
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：泛素-蛋白酶体系统（UPS）和自噬协同维持蛋白质稳态，然而这些通路如何相互作用以调控乳腺癌干细胞（BCSC）仍不清楚。PSMD2是蛋白酶体的一个19S调控亚基，已被认为与多种癌症的不良预后相关，但其在ER+乳腺癌自噬-蛋白稳态网络中的作用尚未明确。在此，我们研究了PSMD2如何通过UPS-自噬串扰影响CSC维持和药物反应，并探讨其抑制如何改变ER+乳腺癌中的自噬调控和CSC脆弱性。
方法：在ER+乳腺癌细胞（MCF-7、ZR-75-1）中构建了稳定PSMD2敲低（ShPSMD2）细胞系。使用羟氯喹（HCQ）调控自噬。Western blotting评估CSC相关标志物（SOX2、OCT4、NANOG）、EMT标志物（E-cadherin、N-cadherin、Vimentin）和自噬蛋白（LC3B-II、p62、p-mTOR）。开展功能实验，包括MTT活力、集落形成、迁移和侵袭，以评估PSMD2依赖性表型。乳腺球和活力实验评估CSC形成和HCQ敏感性。使用NSG小鼠的乳腺脂肪垫和尾静脉异种移植检测肿瘤生长和转移。
结果：PSMD2耗竭降低了CSC频率、乳腺球形成和集落形成，干性标志物（SOX2、OCT4、NANOG）减少，EMT特征逆转，表现为N-cadherin、Vimentin和Snail降低。PSMD2缺失导致LC3B-II和p62积累并伴mTOR抑制，表明自噬周转受损和蛋白稳态破坏。PSMD2缺陷细胞表现出迁移和侵袭减少，且对HCQ诱导的细胞毒性更敏感。HCQ处理进一步以剂量依赖方式降低PSMD2蛋白水平，提示溶酶体应激下蛋白酶体的反馈性去稳定化。这与CD44+/CD24- CSC样群体的丧失和成球能力降低相关。在异种移植中，PSMD2敲低抑制了肿瘤生长以及肺、肝转移，提示PSMD2是连接蛋白稳态与转移进展的关键节点。
结论：PSMD2耗竭扰乱UPS-自噬协调，导致ER+乳腺癌中蛋白稳态丧失和CSC耗竭。这一洞见揭示PSMD2是蛋白水解平衡的关键调控因子，也是基于自噬的治疗策略的潜在靶点。
查看英文原文 English abstract
Background: The ubiquitin-proteasome system (UPS) and autophagy cooperate to maintain protein homeostasis, yet how these pathways interact to regulate breast cancer stem cells (BCSCs) remains unclear. PSMD2, a 19S regulatory subunit of the proteasome, has been linked to poor outcomes in several cancers, but its role within the autophagy-proteostasis network in ER⁺ breast cancer has not been defined. Here, we examined how PSMD2 influences CSC maintenance and drug response through UPS-autophagy crosstalk and explored how its suppression alters autophagy regulation and CSC vulnerability in ER⁺ breast cancer.
Methods: Stable PSMD2 knockdown (ShPSMD2) lines were generated in ER⁺ breast cancer cells (MCF-7, ZR-75-1). Autophagy was modulated using hydroxychloroquine (HCQ). Western blotting assessed CSC-related markers (SOX2, OCT4, NANOG), EMT markers (E-cadherin, N-cadherin, Vimentin), and autophagy proteins (LC3B-II, p62, p-mTOR). Functional assays, including MTT viability, colony formation, migration, and invasion, were performed to evaluate PSMD2-dependent phenotypes. Mammosphere and viability assays assessed CSC formation and HCQ sensitivity. Tumor growth and metastasis were tested using mammary fat-pad and tail-vein xenografts in NSG mice.
Results: PSMD2 depletion reduced CSC frequency, mammosphere formation, and colony formation, with decreased stemness markers (SOX2, OCT4, NANOG) and reversal of EMT features, shown by lower N-cadherin, Vimentin, and Snail. Loss of PSMD2 resulted in LC3B-II and p62 accumulation with mTOR suppression, indicating impaired autophagy turnover and disrupted proteostasis. PSMD2-deficient cells displayed reduced migration and invasion and were more sensitive to HCQ-induced cytotoxicity. HCQ treatment further lowered PSMD2 protein levels in a dose-dependent manner, implying feedback destabilization of the proteasome under lysosomal stress. This correlated with loss of CD44⁺/CD24⁻ CSC-like populations and reduced sphere-forming ability. In xenografts, PSMD2 knockdown suppressed tumor growth and lung, liver metastasis, suggesting that PSMD2 is a critical node linking proteostasis to metastatic progression.
Conclusions: PSMD2 depletion disturbs UPS-autophagy coordination, leading to loss of proteostasis and CSC depletion in ER⁺ breast cancer. This insight reveals PSMD2 as a key regulator of proteolytic balance and a potential target for autophagy-based therapeutic strategies.
利益披露 Disclosure
Y. Lee, None.