journal contribution
posted on 2023-03-31, 04:28 authored by Xinyao Qiu, Shuai Yang, Shan Wang, Jianmin Wu, Bo Zheng, Kaiting Wang, Siyun Shen, Seogsong Jeong, Zhixuan Li, Yanjing Zhu, Tong Wu, Xuan Wu, Rui Wu, Weiwei Liu, Hong-Yang Wang, Lei Chen Supplementary Fig. S1. PD-L1 was regulated by ALKBH5 independent on FTO, METTL3 or METTL14. Supplementary Fig. S2. m6A-seq and mRNA-seq of RBE-shCtrl/shALKBH5 cells. Supplementary Fig. S3. Transient ALKBH5 and its mutant H204A overexpression increased the endogenous ALKBH5 protein level. Supplementary Fig. S4. ALKBH5 deficiency accelerates the degradation of PD-L1 mRNA dependent on YTHDF2. Supplementary Fig. S5. The role of YTHDF2 in regulating PD-L1 expression. Supplementary Fig. S6. Tumor cell intrinsic ALKBH5 facilitates tumor resistance to anti-tumor T cell immunity. Supplementary Fig. S7. T cell expansion was inhibited by tumor-intrinsic ALKBH5. Supplementary Fig. S8. The role of ALKBH5 in the proliferation and migration of ICC cell in vitro. Supplementary Fig. S9. PD-L1 is essential for ALKBH5-regulated anti-tumor T cell immunity. Supplementary Fig. S10. The correlation between ALKBH5 and PD-L1 transcripts in TCGA datasets. Supplementary Fig. S11. ALKBH5 positively correlates with PD-L1 expression in clinical ICC specimens. Supplementary Fig. S12. The role of ALKBH5 in ICC immune microenvironment. Supplementary Fig. S13. ALKBH5 deficiency in ICC cells relates to immunosuppressive microenvironment.
Funding
National Research Program of China
National Natural Science Foundation of China
National Natural Science Foundation of Shanghai
Youth Foundation of Fudan University Shanghai Cancer Center
History
ARTICLE ABSTRACT
N6-methyladenosine (m6A) has been reported as an important mechanism of posttranscriptional regulation. Programmed death-ligand 1 (PD-L1) is a primary immune inhibitory molecule expressed on tumor cells that promotes immune evasion. Here we report ALKBH5 as an important m6A demethylase that orchestrates PD-L1 expression in intrahepatic cholangiocarcinoma (ICC). Regulation of PD-L1 expression by ALKBH5 was confirmed in human ICC cell lines. Sequencing of the m6A methylome identified PD-L1 mRNA as a direct target of m6A modification whose levels were regulated by ALKBH5. Furthermore, ALKBH5 and PD-L1 mRNA were shown to interact. ALKBH5 deficiency enriched m6A modification in the 3′UTR region of PD-L1 mRNA, thereby promoting its degradation in a YTHDF2-dependent manner. In vitro and in vivo, tumor-intrinsic ALKBH5 inhibited the expansion and cytotoxicity of T cells by sustaining tumor cell PD-L1 expression. The ALKBH5-PD-L1–regulating axis was further confirmed in human ICC specimens. Single-cell mass cytometry analysis unveiled a complex role of ALKBH5 in the tumor immune microenvironment by promoting the expression of PD-L1 on monocytes/macrophages and decreasing the infiltration of myeloid-derived suppressor-like cells. Analysis of specimens from patients receiving anti-PD1 immunotherapy suggested that tumors with strong nuclear expression patterns of ALKBH5 are more sensitive to anti-PD1 immunotherapy. Collectively, these results describe a new regulatory mechanism of PD-L1 by mRNA epigenetic modification by ALKBH5 and the potential role of ALKBH5 in immunotherapy response, which might provide insights for cancer immunotherapies.
This study identifies PD-L1 mRNA as a target of ALKBH5 and reveals a role for ALKBH5 in regulating the tumor immune microenvironment and immunotherapy efficacy.
