Department of Thoracic Surgery, Affiliated Hospital of Southwest Medical University, 646000 Luzhou, Sichuan, China.
Front Biosci (Landmark Ed). 2024 Apr 1;29(4):134. doi: 10.31083/j.fbl2904134.
Immune escape is a key factor influencing survival rate of lung adenocarcinoma (LUAD) patients, but molecular mechanism of ubiquitin binding enzyme E2T (UBE2T) affecting immune escape of LUAD remains unclear. The objective was to probe role of UBE2T in LUAD.
Bioinformatics means were adopted for analyzing UBE2T and forkhead box A1 (FOXA1) expression in LUAD tissues, the gene binding sites, the pathway UBE2T regulates, and the correlation between UBE2T and glycolysis genes. Dual luciferase and chromatin immunoprecipitation (ChIP) assays were conducted for validating the binding relationship between the two genes. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot were employed to evaluate UBE2T, FOXA1, and programmed death ligand 1 (PD-L1) levels in cancer cells. MTT assay was conducted for detecting cell viability. Cytotoxicity assay detected CD8+T cell toxicity. Cytokine expression was assayed by enzyme linked immunosorbent assay (ELISA). Extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) were assayed by extracellular flow analyzer. Glycolytic gene expression was analyzed by qRT-PCR, and glycolysis-related indicators were detected by ELISA. Immunohistochemistry (IHC) detected CD8+T cell infiltration in tumor tissues.
FOXA1 and UBE2T were up-regulated in LUAD, and a binding site existed between UBE2T and FOXA1. Overexpressing UBE2T could increase PD-L1 expression and inhibit toxicity of CD8+T cells to LUAD cells. Overexpressing UBE2T repressed CD8+T cell activity in LUAD by activating the glycolysis pathway, and the addition of glycolysis inhibitor 2-deoxy-d-glucose (2-DG) reversed the above results. Mechanistically, FOXA1 promoted the immune escape of LUAD by up-regulating UBE2T and thus mediating glycolysis. experiments revealed that UBE2T knockdown hindered tumor growth, inhibited PD-L1 expression, and facilitated CD8+T cell infiltration.
FOXA1 up-regulated the expression of UBE2T, which activated glycolysis, and thus inhibited activity of CD8+T cells, causing immune escape of LUAD.
免疫逃逸是影响肺腺癌（LUAD）患者生存率的关键因素，但泛素结合酶 E2T（UBE2T）影响 LUAD 免疫逃逸的分子机制尚不清楚。本研究旨在探讨 UBE2T 在 LUAD 中的作用。
采用生物信息学方法分析 LUAD 组织中 UBE2T 和叉头框蛋白 A1（FOXA1）的表达、基因结合位点、UBE2T 调控的通路以及 UBE2T 与糖酵解基因的相关性。通过双荧光素酶和染色质免疫沉淀（ChIP）实验验证这两个基因之间的结合关系。采用定量逆转录聚合酶链反应（qRT-PCR）和蛋白质印迹法检测癌细胞中 UBE2T、FOXA1 和程序性死亡配体 1（PD-L1）的水平。采用 MTT 法检测细胞活力。细胞毒性实验检测 CD8+T 细胞毒性。酶联免疫吸附试验（ELISA）检测细胞因子表达。采用细胞外流量分析仪检测细胞外酸化率（ECAR）和耗氧量（OCR）。qRT-PCR 分析糖酵解基因表达，ELISA 检测糖酵解相关指标。免疫组织化学（IHC）检测肿瘤组织中 CD8+T 细胞浸润。
FOXA1 和 UBE2T 在 LUAD 中上调，UBE2T 和 FOXA1 之间存在结合位点。过表达 UBE2T 可增加 PD-L1 表达并抑制 CD8+T 细胞对 LUAD 细胞的毒性。过表达 UBE2T 通过激活糖酵解通路抑制 LUAD 中 CD8+T 细胞的活性，添加糖酵解抑制剂 2-脱氧-D-葡萄糖（2-DG）可逆转上述结果。机制上，FOXA1 通过上调 UBE2T 介导糖酵解从而促进 LUAD 的免疫逃逸。体内实验表明，UBE2T 敲低可抑制肿瘤生长、降低 PD-L1 表达并促进 CD8+T 细胞浸润。
FOXA1 上调 UBE2T 的表达，激活糖酵解，从而抑制 CD8+T 细胞的活性，导致 LUAD 的免疫逃逸。
