TNFRSF18 Promotes the Proliferation of Breast Cancer Cells by Inhibiting UBL3

Abstract

Objective: The study aims to investigate the role of Glucocorticoid-induced tumor necrosis factor (TNF) receptor superfamily 18 (TNFRSF18, also called GITR) in the proliferation and migration of breast carcinoma cells. Methods: A total of 94 patients with breast carcinoma who underwent surgical excision were enrolled in this study. Cancer tissues and para-cancer tissues were obtained during the operation. Human breast carcinoma cell lines MCF-7 plus MDA-MB-231 were cultivated and transfected. The mRNA or protein levels of TNFRSF18, ubiquitinated-like protein 3 (UBL3), CyclinD1, and matrix metalloproteinase-2 (MMP-2) in breast cancer, para-cancer tissues, MCF-7 cells, and MDA-MB-231 cell lines were quantified using quantitative real-time polymerase chain reaction (qRT-PCR) or western blot analysis. Cell proliferation activity was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. The interaction between TNFRSF18 and UBL3 was verified by immunoluciferase reporter assay. Results: The levels of TNFRSF18 in breast cancer samples were remarkably higher than those in para-cancer samples (p < 0.001). Conversely, the mRNA and protein levels of UBL3 in cancer tissues were significantly lower than those in normal samples (p < 0.001). Both TNFRSF18 and UBL3 mRNA levels were found to be correlated with clinical stage, histological grade, as well as expression levels of the nuclear protein Ki67 and Bromodeoxyuridine (BrdU), respectively. TNFRSF18 overexpression was observed to enhance the proliferation of MDA-MB-231 and MCF-7 cell lines (p < 0.05). Conversely, inhibition of TNFRSF18 or overexpression of UBL3 led to a decrease in the proliferation of MDA-MB-231 and MCF-7 cell lines. Furthermore, an immunoluciferase reporter assay confirmed that UBL3 was a target of TNFRSF18 and was negatively regulated by TNFRSF18. The inhibitory effect of TNFRSF18 downregulation on MDA-MB-231 and MCF-7 cell proliferation was reversed by UBL3 knockdown. Conclusions: The expression of TNFRSF18 is upregulated, while UBL3 expression is downregulated in breast carcinoma samples. TNFRSF18 promotes the proliferation of breast cancer by suppressing UBL3.