Figure 2.
PTPN23 is associated with the PI3K-AKT signaling pathway. (A) Two-way bar chart showing KEGG enrichment analysis of differentially expressed genes identified by RNA-seq after PTPN23 depletion in A375 cells. Upregulated pathways are shown in red, and downregulated pathways in blue. The top 10 pathways are ranked by a normalized enrichment score. Analysis includes n = 3 biological replicates per group. P values were adjusted using the Benjamini and Hochberg approach, with significance thresholds set at P adj ≤0.05 and |log2(fold change)| ≥1 for differential expression. (B) Volcano plots displaying differentially expressed genes in the PI3K-AKT signaling (blue), apoptosis (red), and p53 signaling pathway (violet) after PTPN23 depletion in A375 cells identified by RNA-seq analysis. (C) GSEA of the PI3K-AKT signaling pathway in A375 cells with PTPN23 depletion compared with control cells. (D) Two-way bar chart of KEGG enrichment analysis based on differentially expressed proteins of A375 cells with PTPN23 depletion identified by proteomics. Upregulated proteins are shown in red, and downregulated proteins in blue. Analysis includes n = 3 biological replicates per group. (E) Heatmap of differentially phosphorylated proteins identified by phosphoproteomics after PTPN23 depletion in A375 cells. Upregulated proteins are shown in red, and downregulated proteins in blue. PI3KC2α, AKT2, and AKT3 were highlighted. n = 3 biological replicates per group. P values were adjusted using the Benjamini and Hochberg approach. P adj ≤0.05 and |log2(fold change)| ≥1 were set as the threshold for significantly differential expression. (F) Immunoblot showing phosphorylated and total proteins in A375 and SK-MEL-28 cells after PTPN23 depletion. (G) Immunoblot showing total and phosphorylation levels of indicated proteins in A375 cells after endogenous PTPN23 depletion, with or without Flag-PTPN23 overexpression. (H) mIHC analysis of 44 human melanoma samples with BRAFV600E mutation in a TMA was performed with the indicated antibodies. Representative images of staining are shown. Scale bar, 50 µm. (I) Spearman’s correlation analysis showing the relative expression of PTPN23 and p-AKT2 (S474) in the melanoma TMA samples. (J) Association between the PTPN23 signature score and overall survival in BRAF-mutant and BRAF-WT melanoma patients from TCGA dataset. The PTPN23 signature score was generated based on the top 100 upregulated and 100 downregulated genes from RNA-seq data of control and PTPN23-knockdown cells. Source data are available for this figure: SourceData F2. Refer to the image caption for details.

PTPN23 is associated with the PI3K-AKT signaling pathway. (A) Two-way bar chart showing KEGG enrichment analysis of differentially expressed genes identified by RNA-seq after PTPN23 depletion in A375 cells. Upregulated pathways are shown in red, and downregulated pathways in blue. The top 10 pathways are ranked by a normalized enrichment score. Analysis includes n = 3 biological replicates per group. P values were adjusted using the Benjamini and Hochberg approach, with significance thresholds set at P adj ≤0.05 and |log2(fold change)| ≥1 for differential expression. (B) Volcano plots displaying differentially expressed genes in the PI3K-AKT signaling (blue), apoptosis (red), and p53 signaling pathway (violet) after PTPN23 depletion in A375 cells identified by RNA-seq analysis. (C) GSEA of the PI3K-AKT signaling pathway in A375 cells with PTPN23 depletion compared with control cells. (D) Two-way bar chart of KEGG enrichment analysis based on differentially expressed proteins of A375 cells with PTPN23 depletion identified by proteomics. Upregulated proteins are shown in red, and downregulated proteins in blue. Analysis includes n = 3 biological replicates per group. (E) Heatmap of differentially phosphorylated proteins identified by phosphoproteomics after PTPN23 depletion in A375 cells. Upregulated proteins are shown in red, and downregulated proteins in blue. PI3KC2α, AKT2, and AKT3 were highlighted. n = 3 biological replicates per group. P values were adjusted using the Benjamini and Hochberg approach. P adj ≤0.05 and |log2(fold change)| ≥1 were set as the threshold for significantly differential expression. (F) Immunoblot showing phosphorylated and total proteins in A375 and SK-MEL-28 cells after PTPN23 depletion. (G) Immunoblot showing total and phosphorylation levels of indicated proteins in A375 cells after endogenous PTPN23 depletion, with or without Flag-PTPN23 overexpression. (H) mIHC analysis of 44 human melanoma samples with BRAFV600E mutation in a TMA was performed with the indicated antibodies. Representative images of staining are shown. Scale bar, 50 µm. (I) Spearman’s correlation analysis showing the relative expression of PTPN23 and p-AKT2 (S474) in the melanoma TMA samples. (J) Association between the PTPN23 signature score and overall survival in BRAF-mutant and BRAF-WT melanoma patients from TCGA dataset. The PTPN23 signature score was generated based on the top 100 upregulated and 100 downregulated genes from RNA-seq data of control and PTPN23-knockdown cells. Source data are available for this figure: SourceData F2.

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