claudin-1在口腔鳞癌SCCKN细胞表达的分子机制分析

IF 0.4 Q4 DENTISTRY, ORAL SURGERY & MEDICINE

Journal of Oral and Maxillofacial Surgery Medicine and Pathology Pub Date : 2024-12-29 DOI:10.1016/j.ajoms.2024.12.017

Tadayoshi Nobumoto , Sachiko Yamasaki , Atsuko Hamada , Mirai Higaki , Nanako Ito , Fumitaka Obayashi , Yasutaka Ishida , Tomoaki Hamana , Tomoaki Shintani , Ryouji Tani , Koichi Koizumi , Souichi Yanamoto , Yasutaka Hayashido

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引用次数: 0

摘要

目的p53泛素连接酶人双分钟2 （HDM2）的表达增加了β-catenin的核易位和紧密连接主要组成蛋白CLD1的表达。麻木蛋白X1配体（LNX1）是一种E3泛素连接酶，在犬肾小管上皮细胞中与CLD1结合并促进其被溶酶体降解。然而，LNX1对口腔鳞状细胞癌（OSCC） CLD1表达的影响尚不清楚。因此，在本研究中，我们旨在阐明LNX1影响OSCC细胞CLD1表达的分子机制。方法采用共免疫沉淀法和哺乳动物双杂交法，研究β-catenin/TCF通路在OSCC SCCKN细胞CLD1表达中的作用，探讨LNX1与CLD1表达的关系。此外，我们评估了HDM2对SCCKN细胞中LNX1表达的影响。结果SCCKN细胞通过β-catenin通路调控CLD1表达，表明HDM2上调时β-catenin的核易位升高了CLD1水平。抑制剂实验显示SCCKN细胞中的CLD1蛋白被蛋白酶体或溶酶体降解。联合免疫沉淀和哺乳动物双杂交实验表明，CLD1在SCCKN细胞中与LNX1结合。此外，在SCCKN细胞中，LNX1p80与HDM2结合并发生蛋白酶体或溶酶体降解。结论hdm2诱导的LNX1p80降解的增加抑制了蛋白酶体和溶酶体对CLD1的降解，从而增加了SCCKN细胞中CLD1的水平。

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Analysis of the molecular mechanism of claudin-1 expression in SCCKN cells, oral squamous cell carcinoma cell line

Objectives

Expression of human double minute-2 (HDM2), a p53 ubiquitin ligase, increases the nuclear translocation of β-catenin and expression of claudin-1 (CLD1), a major component protein of tight junctions. Ligand of numb protein X1 (LNX1), an E3 ubiquitin ligase, binds to CLD1 and promotes its degradation by lysosomes in canine renal tubular epithelial cells. However, effect of LNX1 on CLD1 expression in oral squamous cell carcinoma (OSCC) remains unknown. Therefore, in this study, we aimed to elucidate the molecular mechanisms by which LNX1 affects CLD1 expression in OSCC cells.

Methods

We investigated the role of the β-catenin/TCF pathway in CLD1 expression to determine the relationship between LNX1 and CLD1 expression in SCCKN cell line, an OSCC cell line, via co-immunoprecipitation and mammalian two-hybrid assays. Additionally, we evaluated the effect of HDM2 on LNX1 expression in SCCKN cells.

Results

CLD1 expression was regulated via the β-catenin pathway, indicating that nuclear translocation of β-catenin upon HDM2 upregulation elevated the CLD1 levels in SCCKN cells. Inhibitor assays revealed that CLD1 protein in SCCKN cells was degraded by proteasomes or lysosomes. Co-immunoprecipitation and mammalian two-hybrid assays showed that CLD1 bound to LNX1 in SCCKN cells. Furthermore, LNX1p80 bound to HDM2 and underwent proteasomal or lysosomal degradation in SCCKN cells.

Conclusions

Our results suggest that HDM2-induced increase in LNX1p80 degradation suppresses CLD1 degradation by the proteasomes and lysosomes, thereby increasing the CLD1 levels in SCCKN cells.

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