Deciphering the Divergent Gene Expression Landscapes of m6A/m5C/m1A Methylation Regulators in Hepatocellular Carcinoma Through Single-Cell and Bulk RNA Transcriptomic Analysis

Figures & data

Figure 1 Dysregulated m6A/m5C/m1A genes identified from the bulk sequencing of The Cancer Genome Atlas (TCGA) cohort. *Indicates a significant change (p<0.05) between the hepatocellular carcinoma (HCC) and the adjacent para-cancerous liver tissue.

Figure 2 The clustered heatmap of the dysregulated m6A/m5C/m1A genes in HCC and in adjacent para-cancerous liver tissue.

Table 1 The Dysregulated m6A/m5C/m1A Genes Identified from the scRNA-Seq in Different Cell Types in the HCC vs the Adjacent Para-Cancerous Liver Tissue

Symbols	Gene Name	Log2 Fold Change	Adj. p-value	Cell Type	Methylation Type
ALKBH5	AlkB Homolog 5, RNA Demethylase	−1.33	<0.00001	CD8+ T-cells	Erasers_m6a
ALKBH5	AlkB Homolog 5, RNA Demethylase	−1.82	0.000028	NK cells	Erasers_m6a
DNMT1	DNA Methyltransferase 1	2.57	<0.00001	Hepatocytes	Writers_m5c
DNMT3A	DNA Methyltransferase 3 Alpha	−1.49	0.006008	NK cells	Writers_m5c
IGF2BP2	Insulin Like Growth Factor 2 MRNA Binding Protein 2	−2.95	<0.00001	Endothelial cells	Readers_m6a
METTL3	Methyltransferase 3, N6-Adenosine-Methyltransferase Complex Catalytic Subunit	1.02	0.002054	Endothelial cells	Writers_m6a
NSUN5	NOP2/Sun RNA Methyltransferase 5	1.35	0.0009564	Hepatocytes	Writers_m5c
PRRC2A	Proline Rich Coiled-Coil 2A	−1.25	0.0277234	NK cells	Readers_m6a
RBM15B	RNA Binding Motif Protein 15B	−1.06	<0.00001	CD8+ T-cells	Writers_m6a
TRMT6	TRNA Methyltransferase 6 Non-Catalytic Subunit	1.01	0.027211	Hepatocytes	Writers_m1a
WTAP	WT1 Associated Protein	−1.23	<0.00001	NK cells	Writers_m6a
YBX1	Y-Box Binding Protein 1	−1.01	<0.00001	CD8+ T-cells	Readers_m5c
YBX1	Y-Box Binding Protein 1	−1.24	<0.00001	NK cells	Readers_m5c
YTHDC1	YTH N6-Methyladenosine RNA Binding Protein C1	−1.01	0.000043	Hepatocytes	Reader_m6a
YTHDC1	YTH N6-Methyladenosine RNA Binding Protein C1	−1.01	0.000043	Hepatocytes	Readers_m6a
IGF2BP1	Insulin Like Growth Factor 2 MRNA Binding Protein 1	Inf	0.036279	Hepatocytes	Readers_m6a
IGF2BP3	Insulin Like Growth Factor 2 MRNA Binding Protein 3	Inf	<0.00001	Hepatocytes	Readers_m6a

Figure 3 Single-cell RNA-sequencing analysis identified based on the HCC vs adjacent para-cancerous liver tissue, GSE categories, cell cycles, and major clusters of the cells.

Figure 4 (A) The bubble plot conducted to visualize the distribution of m6A/m5C/m1A genes among different cell types. To evaluate the statistical significance of each interaction score, a one-sided Wilcoxon rank-sum test was employed. (B) The river plot shows the association of dysregulated m6A/m5C/m1A genes to different cell types. GSE, Gene Series Accession; UMAP, Uniform Manifold Approximation and Projection; G1, Gap 1; S, Synthesis of DNA; G2 M, Gap 2 and Mitosis. (C) Summary of the expression of scRNA-seq-identified dysregulated m6A/m5C/m1A genes in the bulk sequencing from TCGA cohort. (D) The clustered heatmap of these dysregulated m6A/m5C/m1A genes in HCC and in adjacent para-cancerous liver tissue according to the bulk sequencing from TCGA cohort. *Indicates a significant change (p<0.05) between the hepatocellular carcinoma (HCC) and the adjacent para-cancerous liver tissue.

Figure 5 (A) A real-time PCR was performed to measure the expression of dysregulated m6A/m5C/m1A genes identified from the scRNA-seq. (B) A real-time PCR was performed to measure the expression of dysregulated m6A/m5C/m1A genes identified from selected representative dysregulated m6A/m5C/m1A genes from TCGA. (C) Kaplan–Meier (K–M) survival analysis of six dysregulated methylation-related genes using Timer2.0. *Indicates a significant change (p<0.05) between the hepatocellular carcinoma (HCC) and the adjacent para-cancerous liver tissue.

Figure 6 Correlation of dysregulated m6A/m5C/m1A genes with immune cell infiltration using Timer2.0. *Indicates a significant change (p<0.05) of the type of infiltrated cells between the hepatocellular carcinoma (HCC) and the adjacent para-cancerous liver tissue.