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Fig. 1 | Experimental Hematology & Oncology

Fig. 1

From: Secretogranin II influences the assembly and function of MHC class I in melanoma

Fig. 1

SCG2 is more strongly expressed in melanoma compared to healthy skin and reduces the overall survival (OS) of melanoma patients (A) SCG2 expression data from DFCI, Nature Medicine 2019. Kaplan–Meier curve showing OS of patients (n = 121) with metastatic melanoma with high intratumoral SCG2 expression (Log2 SCG2 ≥ 1) compared to patients with low intratumoral SCG2 expression (Log2 SCG2 < 1). B Patient data obtained from the GSE7553 database showing SCG2 expression levels as log2 in normal skin (n = 5), primary melanoma (n = 14), and melanoma metastases (n = 40). Statistical analysis was conducted using one-way ANOVA. C SCG2 immunohistochemistry (IHC) staining of patient samples from nevi (n = 16), primary melanoma (n = 37), and melanoma metastases (n = 52). Statistical analysis was conducted using one-way ANOVA. (D) Confirmation and quantification of SCG2 overexpression (OE) in WM266-4 and C32 melanoma cells on mRNA (upper panel) and protein (lower panel) level. Empty vector (EV) cells were used as a reference. 18S was used as an internal control. GAPDH was used as loading control. Data represent mean ± s.e.m. (n ≥ 3) (E) Cell cycle analysis of WM266-4 (left panel) and C32 (right panel) EV and SCG2 OE cells. DNA was stained using propidium iodide (PI) and the number of PI-positive cells was determined using flow cytometry (n = 3). F Fold change of mRNA expression levels of the ER markers and APM components calreticulin (CALR) and calnexin (CANX) in WM266-4 (left panel) and C32 (right panel) SCG2 OE cells compared to EV control (ctrl). Data represent mean ± s.e.m. (n ≥ 3). G Fold change of mRNA expression of the APM components TAP1, TAP2, B2M, and TAPBP (tapasin) in SCG2 OE cells (WM266-4, left panel, and C32, right panel) compared to EV control. 18S was used as endogenous control. Data represent mean ± s.e.m. (n ≥ 3). H Protein levels of the APM components calnexin, TAP2, TAP1, calreticulin, tapasin, and B2M in WM266-4 (left panel) and C32 (right panel) EV and SCG2 OE cells. GAPDH was used as loading control. I Correlation of the expression of SCG2 and HLA-A, HLA-B, and HLA-C, respectively, in melanoma patients (n = 87) according to the data from GSE7553. J Mean fluorescence intensity (MFI) of HLA-ABC-positive ( +) WM266-4 (left panel) and C32 (right panel) cells comparing SCG2 OE to EV control. Data represent mean ± s.e.m. (n ≥ 3). K T cell cytotoxicity assay performed with MART-1-specific T cells measured by xCELLigence RTCA impedance assay. The interaction of the WM266-4 (left panel) and C32 (right panel) cells with the gold biosensors was measured through the cellular impedance. This impedance value is plotted as normalized cell index, which correlates with the cell number. An increase of the normalized cell index indicates cell proliferation while a decrease represents the neutralization of melanoma cells through T cell-mediated cytotoxicity. We compared the normalized cell index of WM266-4 (left panel) and C32 (right panel) EV (black) and SCG2 OE (red) cells over time. Data represent mean ± s.e.m. of three independent experiments (n = 3). *p < 0.05; **p < 0.01; ***p < 0.001; “ns” refers to p ≥ 0.05

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