Abstract
Chronic inflammation is a well-established risk factor in the development of gastric cancer (GC). Tumor necrosis factor α-induced protein 3 (TNFAIP3, also known as A20) is an inflammation-associated protein that functions as an oncogene in various cancers, but the role of A20 in GC progression remains unclear. In this study, clinical analyses revealed that elevated A20 expression in GC patients was significantly associated with increased tumor aggressiveness and metastatic potential. Functionally, A20 overexpression enhanced GC cell migration, whereas its knockdown suppressed this effect. Moreover, A20 promoted epithelial-mesenchymal transition and reduced the tight junction protein occludin. Mechanistically, A20 induced occludin endocytosis and lysosomal degradation via its ovarian tumor (OTU) domain. Pull-down assays revealed that A20 interacts with the migration-related protein RhoA, increasing its stability and thereby sustaining ROCK2 phosphorylation, which contributes to occludin degradation. PLA further showed that mutation of the OTU domain disrupted the interaction between A20 and RhoA in AGS cells, indicating the necessity of the OTU domain for this interaction. In conclusion, our findings demonstrate that A20 promotes GC cell migration by stabilizing RhoA and facilitating occludin degradation, underscoring A20 as a potential therapeutic target to inhibit GC metastasis.
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References
Siegel RL, Miller KD, Fuchs HE, Jemal A. Cancer Statistics, 2021. CA: A Cancer J Clin. 2021;71:7–33.
Kinami S, Saito H, Takamura H. Significance of lymph node metastasis in the treatment of gastric cancer and current challenges in determining the extent of metastasis. Front Oncol. 2021;11:806162.
Guan WL, He Y, Xu RH. Gastric cancer treatment: recent progress and future perspectives. J Hematol Oncol. 2023;16:57.
Hibino S, Kawazoe T, Kasahara H, Itoh S, Ishimoto T, Sakata-Yanagimoto M. et al. Inflammation-induced tumorigenesis and metastasis. Int J Mol Sci. 2021;22:5421.
O’Brien VP, Kang Y, Shenoy MK, Finak G, Young WC, Dubrulle J, et al. Single-cell profiling uncovers a muc4-expressing metaplastic gastric cell type sustained by helicobacter pylori-driven inflammation. Cancer Res Commun. 2023;3:1756–69.
Rihawi K, Ricci AD, Rizzo A, Brocchi S, Marasco G, Pastore LV. et al. Tumor-associated macrophages and inflammatory microenvironment in gastric cancer: novel translational implications. Int J Mol Sci. 2021;22:3805.
Waldum H, Fossmark R. Inflammation and digestive cancer. Int J Mol Sci. 2023;24:13503.
Krikos A, Laherty CD, Dixit VM. Transcriptional activation of the tumor necrosis factor alpha-inducible zinc finger protein, A20, is mediated by kappa B elements. J Biol Chem. 1992;267:17971–6.
Mothes J, Ipenberg I, Arslan S, Benary U, Scheidereit C, Wolf J. A quantitative modular modeling approach reveals the effects of different A20 feedback implementations for the NF-kB signaling dynamics. Front Physiol. 2020;11:896.
Peng X, Zhang C, Zhou ZM, Wang K, Gao JW, Qian ZY, et al. A20 attenuates pyroptosis and apoptosis in nucleus pulposus cells via promoting mitophagy and stabilizing mitochondrial dynamics. Inflamm Res. 2022;71:695–710.
Luo M, Wang X, Wu S, Yang C, Su Q, Huang L, et al. A20 promotes colorectal cancer immune evasion by upregulating STC1 expression to block “eat-me” signal. Signal Transduct Target Ther. 2023;8:312.
Paul S, Bhardwaj M., Kang SC. GSTO1 confers drug resistance in HCT-116 colon cancer cells through an interaction with TNFalphaIP3/A20. Int J Mol Sci. 2022;61:136.
Liu J, Yang S, Wang Z, Chen X, Zhang Z. Ubiquitin ligase A20 regulates p53 protein in human colon epithelial cells. J Biomed Sci. 2013;20:74.
Nehme Z, Roehlen N, Dhawan P. Baumert TF: tight junction protein signaling and cancer biology. Cells. 2023;12:243.
Bergers G, Fendt SM. The metabolism of cancer cells during metastasis. Nat Rev Cancer. 2021;21:162–80.
Huang Y, Hong W, Wei X. The molecular mechanisms and therapeutic strategies of EMT in tumor progression and metastasis. J Hematol Oncol. 2022;15:129.
Attiq A, Afzal S. Trinity of inflammation, innate immune cells and cross-talk of signalling pathways in tumour microenvironment. Front Pharmacol. 2023;14:1255727.
Ma J, Wang H, Guo S, Yi X, Zhao T, Liu Y, et al. A20 promotes melanoma progression via the activation of the Akt pathway. Cell Death Dis. 2020;11:794.
Lee JH, Jung SM, Yang KM, Bae E, Ahn SG, Park JS, et al. A20 promotes metastasis of aggressive basal-like breast cancers through multi-monoubiquitylation of Snail1. Nat Cell Biol. 2017;19:1260–73.
Du B, Liu M, Li C, Geng X, Zhang X, Ning D, et al. The potential role of TNFAIP3 in malignant transformation of gastric carcinoma. Pathol, Res Pract. 2019;215:152471.
Wisnieski F, Santos LC, Calcagno DQ, Geraldis JC, Gigek CO, Anauate AC, et al. The impact of DNA demethylation on the upregulation of the NRN1 and TNFAIP3 genes associated with advanced gastric cancer. J Mol Med. 2020;98:707–17.
Osanai M, Murata M, Nishikiori N, Chiba H, Kojima T, Sawada N. Epigenetic silencing of occludin promotes tumorigenic and metastatic properties of cancer cells via modulations of unique sets of apoptosis-associated genes. Cancer Res. 2006;66:9125–33.
Wang M, Liu Y, Qian X, Wei N, Tang Y, Yang J. Downregulation of occludin affects the proliferation, apoptosis and metastatic properties of human lung carcinoma. Oncol Rep. 2018;40:454–62.
Li Y, Yuan T, Zhang H, Liu S, Lun J, Guo J, et al. PHD3 inhibits colon cancer cell metastasis through the occludin-p38 pathway. Acta Biochim Biophys Sin. 2023;55:1749–57.
Mortell KH, Marmorstein AD, Cramer EB. Fetal bovine serum and other sera used in tissue culture increase epithelial permeability. In Vitro Cell Dev Biol. 1993;29A:235–8.
Kim KA, Kim D, Kim JH, Shin YJ, Kim ES, Akram M, et al. Autophagy-mediated occludin degradation contributes to blood-brain barrier disruption during ischemia in bEnd.3 brain endothelial cells and rat ischemic stroke models. Fluids Barriers CNS. 2020;17:21.
Utech M, Mennigen R, Bruewer M. Endocytosis and recycling of tight junction proteins in inflammation. J Biomed Biotechnol. 2010;2010:484987.
Soni D, Wang DM, Regmi SC, Mittal M, Vogel SM, Schluter D, et al. Deubiquitinase function of A20 maintains and repairs endothelial barrier after lung vascular injury. Cell Death Discov. 2018;4:60.
Yin H, Karayel O, Chao YY, Seeholzer T, Hamp I, Plettenburg O, et al. A20 and ABIN-1 cooperate in balancing CBM complex-triggered NF-kappaB signaling in activated T cells. Cell Mol Life Sci. 2022;79:112.
Wang Y, Zhang J, Yi XJ, Yu FS. Activation of ERK1/2 MAP kinase pathway induces tight junction disruption in human corneal epithelial cells. Exp Eye Res. 2004;78:125–36.
Feng S, Zou L, Wang H, He R, Liu K., Zhu H. RhoA/ROCK-2 pathway inhibition and tight junction protein upregulation by catalpol suppresses lipopolysaccharide-induced disruption of blood-brain barrier permeability. Molecules. 2018;23:2371.
Martin TA, Jiang WG. Loss of tight junction barrier function and its role in cancer metastasis. Biochim Biophys Acta. 2009;1788:872–91.
Saitou M, Furuse M, Sasaki H, Schulzke JD, Fromm M, Takano H, et al. Complex phenotype of mice lacking occludin, a component of tight junction strands. Mol Biol Cell. 2000;11:4131–42.
Wroblewski LE, Shen L, Ogden S, Romero-Gallo J, Lapierre LA, Israel DA, et al. Helicobacter pylori dysregulation of gastric epithelial tight junctions by urease-mediated myosin II activation. Gastroenterology. 2009;136:236–46.
Huang S, Zhang J, Li Y, Xu Y, Jia H, An L, et al. Downregulation of Claudin5 promotes malignant progression and radioresistance through Beclin1-mediated autophagy in esophageal squamous cell carcinoma. J Transl Med. 2023;21:379.
Kolodziej LE, Lodolce JP, Chang JE, Schneider JR, Grimm WA, Bartulis SJ, et al. TNFAIP3 maintains intestinal barrier function and supports epithelial cell tight junctions. PloS one. 2011;6:e26352.
Yokoyama Y, Tamachi T, Iwata A, Maezawa Y, Meguro K, Yokota M, et al. A20 (Tnfaip3) expressed in CD4(+) T cells suppresses Th2 cell-mediated allergic airway inflammation in mice. Biochem Biophys Res Commun. 2022;629:47–53.
Li L, Hailey DW, Soetandyo N, Li W, Lippincott-Schwartz J, Shu HB, et al. Localization of A20 to a lysosome-associated compartment and its role in NFkappaB signaling. Biochim Biophys Acta. 2008;1783:1140–9.
Glowacka WK, Alberts P, Ouchida R, Wang JY, Rotin D. LAPTM5 protein is a positive regulator of proinflammatory signaling pathways in macrophages. J Biol Chem. 2012;287:27691–702.
Sharif M, Baek YB, Naveed A, Stalin N, Kang MI, Park SI. Porcine sapovirus-induced tight junction dissociation via activation of RhoA/ROCK/MLC signaling pathway. J Virol. 2021;95:e00051–21.
Hasan MK, Widhopf GF 2nd, Zhang S, Lam SM, Shen Z, et al. Wnt5a induces ROR1 to recruit cortactin to promote breast-cancer migration and metastasis. NPJ breast cancer. 2019;5:35.
Xu Z, Gu C, Yao X, Guo W, Wang H, Lin T, et al. CD73 promotes tumor metastasis by modulating RICS/RhoA signaling and EMT in gastric cancer. Cell Death Dis. 2020;11:202.
Terry S, Nie M, Matter K, Balda MS. Rho signaling and tight junction functions. Physiology. 2010;25:16–26.
Kim JH, Park S, Lim SM, Eom HJ, Balch C, Lee J, et al. Rational design of small molecule RHOA inhibitors for gastric cancer. Pharmacogenomics J. 2020;20:601–12.
Nam S, Lee Y, Kim JH. RHOA protein expression correlates with clinical features in gastric cancer: a systematic review and meta-analysis. BMC Cancer. 2022;22:798.
Chen Z, Liu S, Xia Y, Wu K. MiR-31 regulates Rho-associated kinase-myosin light chain (ROCK-MLC) pathway and inhibits gastric cancer invasion: roles of RhoA. Med Sci Monit. 2016;22:4679–91.
Oh M, Batty S, Banerjee N, Kim TH. High extracellular glucose promotes cell motility by modulating cell deformability and contractility via the cAMP-RhoA-ROCK axis in human breast cancer cells. Mol Biol Cell. 2023;34:ar79.
Xie L, Huang H, Zheng Z, Yang Q, Wang S, Chen Y, et al. MYO1B enhances colorectal cancer metastasis by promoting the F-actin rearrangement and focal adhesion assembly via RhoA/ROCK/FAK signaling. Ann Transl Med. 2021;9:1543.
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This study was supported by the Taiwan Ministry of Science and Technology (NSTC 109-2314-B-006-028-MY2).
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YTK: Methodology, investigation, visualization, writing-original draft. HCW: Conceptualization, writing-review and editing. YSS: Conceptualization, resources, supervision, funding acquisition, project administration, writing-review and editing.
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All procedures involving human tissues and data in this study were approved by the Institutional Review Board (IRB) of NCKUH (IRB No. B-ER-107-432). The requirement for informed consent was waived by the IRB. All the animal experiments were conducted in compliance with national legislation and were approved by the Institutional Animal Care and Use Committee of National Cheng Kung University, under approval numbers IACUC/NCKU/108226 and IACUC/NCKU/112301.
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Kuo, YT., Wang, HC. & Shan, YS. A20 enhances the migration and metastasis of gastric cancer cells by promoting occludin degradation. Cell Death Discov. (2026). https://doi.org/10.1038/s41420-026-03082-2
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DOI: https://doi.org/10.1038/s41420-026-03082-2
