American Association for Cancer Research
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Figure S1-S7 from Skp2-Mediated Stabilization of MTH1 Promotes Survival of Melanoma Cells upon Oxidative Stress

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journal contribution
posted on 2023-03-31, 01:00 authored by Jia Yu Wang, Guang Zhi Liu, James S. Wilmott, Ting La, Yu Chen Feng, Hamed Yari, Xu Guang Yan, Rick F. Thorne, Richard A. Scolyer, Xu Dong Zhang, Lei Jin

Figure S1 shows the turnover rates of MTH1 mRNA remained similar between melanoma cells with relatively high (Mel-FH) and low (IgR3) expression of the transcript. Figure S2 shows silencing or overexpression of Skp2 did not alter the expression levels of the MTH1 transcript in melanoma cells. Figure S3 shows silencing of MTH1 did not impinge on the expression Skp2. Figure S4 shows Skp2-NES retains the ability to bind to MTH1. Figure S5 shows Skp2 silencing inhibits melanoma cell proliferation but does trigger apoptosis. Figure S6 shows the oxidative stress inducer elesclomol increases ROS levels. Figure S7 shows BRAF mutant melanoma cells selected for resistance to vemurafenib are more susceptible to killing by knockdown of Skp2 or MTH1.


National Health and Medical Research Council

Cancer Council NSW



MTH1 helps prevent misincorporation of ROS-damaged dNTPs into genomic DNA; however, there is little understanding of how MTH1 itself is regulated. Here, we report that MTH1 is regulated by polyubiquitination mediated by the E3 ligase Skp2. In melanoma cells, MTH1 was upregulated commonly mainly due to its improved stability caused by K63-linked polyubiquitination. Although Skp2 along with other components of the Skp1-Cullin-F-box (SCF) ubiquitin ligase complex was physically associated with MTH1, blocking the SCF function ablated MTH1 ubiquitination and expression. Conversely, overexpressing Skp2-elevated levels of MTH1 associated with an increase in its K63-linked ubiquitination. In melanoma cell lines and patient specimens, we observed a positive correlation of Skp2 and MTH1 expression. Mechanistic investigations showed that Skp2 limited DNA damage and apoptosis triggered by oxidative stress and that MAPK upregulated Skp2 and MTH1 to render cells more resistant to such stress. Collectively, our findings identify Skp2-mediated K63-linked polyubiquitination as a critical regulatory mechanism responsible for MTH1 upregulation in melanoma, with potential implications to target the MAPK/Skp2/MTH1 pathway to improve its treatment. Cancer Res; 77(22); 6226–39. ©2017 AACR.