Shen L, Zhang D, Gao S
Henan Provincial Key Laboratory of Cancer Epigenetics, Cancer Institute, First Affiliated Hospital, College of Clinical Medicine, Henan University of Science and Technology, Luoyang 471003, China.
The 989th Hospital of the People's Liberation Army Joint Service Support Force, Luoyang 471003, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2023 Jul 20;43(7):1155-1163. doi: 10.12122/j.issn.1673-4254.2023.07.12.
To investigate the effect of (Pg) infection on IFNGR1 palmitoylation and biological behaviors of esophageal squamous cell carcinoma (ESCC) cells and the clinical implications.
The expression levels of IFNGR1 protein in ESCC cell lines KYSE30 and KYSE70 were detected using Western blotting at 24 and 48 h after Pg infection, and 2-BP was used to detect IFNGR1 palmitoylation in the cells. KYSE70 cells with wild-type IFNGR1 (IFNGR1-WT cells) and with IFNGR1-C122A palmitoylation site mutation induced by site-specific mutagenesis (IFNGR1-C122A cells) were both infected with Pg, and the changes in palmitoylation of IFNGR1-C122A were analyzed using immunofluorescence and Click-iT assays. The changes in proliferation, migration and invasion ability of the infected cells were evaluated using plate cloning assay, scratch assay and Transwell assay, and IFNGR1 co-localization with lysosomal marker LAMP2 was dected using immunofluorescence assay. Immunohistochemistry was used to detect Pg infection and IFNGR1 protein expression in 50 ESCC tissues, and their correlation with the clinicopathological characteristics and survival outcomes of the patients was analyzed.
Pg infection down-regulated the protein expression of IFNGR1 in ESCC and promoted IFNGR1 palmitoylation at site 122. In IFNGR1-WT cells, Pg infection significantly enhanced cell proliferation, migration and invasion ( < 0.05). Similarly, Pg also significantly promoted proliferation, migration and invasion of IFNGR1-C122A cells, but to a lesser extent as compared with the wild-type cells ( < 0.05). Immunofluorescence assay showed that Pg and ZDHHC3 promoted IFNGR1 degradation within the lysosome. Immunohistochemical studies of the ESCC tissue samples showed a negative correlation between IFNGR1 and Pg expression, and a reduced IFNGR1 expression was correlated with a poorer survival outcome of the patient.
Pg infection enhances IFNGR1 palmitoylation to promote progression of ESCC, and elimination of Pg and inhibiting IFNGR1 palmitoylation may effectively control ESCC progression.
探讨牙龈卟啉单胞菌（Pg）感染对食管鳞状细胞癌（ESCC）细胞中IFNGR1棕榈酰化及生物学行为的影响及其临床意义。
采用蛋白质免疫印迹法检测Pg感染24小时和48小时后ESCC细胞系KYSE30和KYSE70中IFNGR1蛋白的表达水平，并用2-溴棕榈酸检测细胞中IFNGR1的棕榈酰化。将野生型IFNGR1的KYSE70细胞（IFNGR1-WT细胞）和经定点诱变诱导的IFNGR1-C122A棕榈酰化位点突变的细胞（IFNGR1-C122A细胞）均感染Pg，采用免疫荧光和Click-iT检测法分析IFNGR1-C122A棕榈酰化的变化。采用平板克隆实验、划痕实验和Transwell实验评估感染细胞增殖、迁移和侵袭能力的变化，采用免疫荧光实验检测IFNGR1与溶酶体标志物LAMP2的共定位。采用免疫组织化学法检测50例ESCC组织中Pg感染及IFNGR1蛋白表达情况，并分析其与患者临床病理特征及生存结局的相关性。
Pg感染下调ESCC中IFNGR1蛋白表达并促进122位点的IFNGR1棕榈酰化。在IFNGR1-WT细胞中，Pg感染显著增强细胞增殖、迁移和侵袭能力（P<0.05）。同样，Pg也显著促进IFNGR1-C122A细胞的增殖、迁移和侵袭，但与野生型细胞相比程度较小（P<0.05）。免疫荧光实验显示，Pg和ZDHHC3促进IFNGR1在溶酶体内降解。ESCC组织样本的免疫组织化学研究显示，IFNGR1与Pg表达呈负相关，IFNGR1表达降低与患者较差的生存结局相关。
Pg感染增强IFNGR1棕榈酰化以促进ESCC进展，清除Pg并抑制IFNGR1棕榈酰化可能有效控制ESCC进展。
