Analysis at the single-cell level indicates an important role of heterogeneous global DNA methylation status on the progression of lung adenocarcinoma

Quan-Fang Chen,Qing-Yun Pan,Ying-Ju Wang,Han Gao,Xiao-Ning Zhong

doi:10.1038/s41598-021-02786-y

Abstract

Aberrant DNA modifications affect the tumorigenesis and progression of lung cancer. However, the global methylation status of tumor cells and the heterogeneous methylation status of cells within the same tumor need further study. We used publicly available single-cell RNAseq data to investigate the impact and diversity of global methylation status on lung adenocarcinoma. Clustering cells into subgroups and cell differentiation pseudotime analysis, based on expression profile, demonstrated that the global methylation status was crucial to lung adenocarcinoma function and progression. Hypermethylated tumor cells had increased activity related to the hypoxia response. Hyper- and hypomethylated cells indicated upregulation in pathways involving focal adhesion and cell junctions. Pseudotime analysis showed that cell clusters with unique methylation activities were located at the ends of the putative trajectories, suggesting that DNA methylation and demethylation activities were essential to tumor cell progression. Expression of SPP1 was associated with the global methylation status of tumor cells and with patient prognosis. Our study identified the importance and diversity of global DNA methylation status by analysis at the single-cell level. Our findings provide new information about the global DNA methylation status of tumor cells and suggest new approaches for precision medical treatments for lung adenocarcinoma.

Highlights

Lung cancer is a commonly diagnosed cancer that is a major cause of cancer deaths worldwide
We used the Seurat pipeline (Fig. 1) to characterize the detailed DNA methylation status of 6251 tumor cells from 3 patients with lung adenocarcinomas at the core, middle, and border sites into eight clusters (Fig. 2A). Analysis of these data using t-distributed stochastic neighbor embedding (t-SNE) plots according to sample site showed that cells from cluster-6 were all from the core region of the tumor, cells from cluster-4 were enriched in the core region, and cells from cluster-5 were depleted from the middle region (Fig. 2B)
DNA methylation is mainly catalyzed by enzymes in the DNA methyltransferase (DNMT) family, including DNMT3A and DNMT3B and DNMT17

Summary

Introduction

Lung cancer is a commonly diagnosed cancer that is a major cause of cancer deaths worldwide. Previous studies of the biological role of DNA methylation in lung cancer mainly focused on single genes, such as the relationship of the methylation of a promoter of a specific gene with its expression, and the relationship of the expression of this specific gene with oncogenesis and tumor progression. Despite extensive research on the hypermethylation of gene promoters, few studies have examined the genome-wide methylation activity of lung cancer cells. The development of single-cell sequencing technology made it possible to profile global methylation levels in lung cancer at the resolution of single cells. We used single-cell sequencing data from 3 patients with lung adenocarcinomas, and compared expression profiles of samples from the core, middle, and border of the tumor. We used single-cell pseudotime analysis to verify the role of global methylation on the progression of lung adenocarcinoma

Methods

Results

Conclusion