SERP1 is a novel marker of poor prognosis in pancreatic ductal adenocarcinoma patients via anti-apoptosis and regulating SRPRB/NF-κB axis

Abstract

Stress associated endoplasmic reticulum protein 1 (SERP1), can cause accumulation of unfolded proteins in ER stress. However, studies on the role of SERP1 in pancreatic ductal adenocarcinoma (PDAC) are still incomplete. The present study aimed at identifying whether SERP1 acts as a potential novel prognostic marker of PDAC, and analyzed its possible mechanism. GEO database analysis showed SERP1 was significantly upregulated in PDAC tissues, and strongly associated with advanced clinical stage of PDAC patients from TCGA database. Univariate and multivariate Cox regression analysis further revealed SERP1 high expression was an independent factor for the prognosis of PDAC. Gene set enrichment analysis (GSEA) revealed that SERP1 was mainly involved in regulating cell apoptosis and nuclear factor-κB (NF-κB) signaling pathway, and downregulated SERP1 significantly promoted PANC-1 cell apoptosis. To further explore its possible mechanism, protein-protein interaction (PPI) and gene ontology (GO) analysis showed the functions of proteins interacting with SERP1 were mainly enriched in regulating cell apoptosis, and SRP receptor β subunit (SRPRB) was the core of the whole PPI network. The expression of SERP1 was negatively correlated with SRPRB expression. In vitro, downregulated SERP1 significantly increased SRPRB expression. Furthermore, upregulated SRPRB could increase cell apoptosis rate and decreased the expression level of NF-κB and the phosphorylation NF-κB. The above results indicated that SERP1 as a potential novel prognostic marker of PDAC probably via regulating cell apoptosis and NF-κB activation, which may be associated with SRPRB.

Figure 1

Flow chart of the derivation and function analysis of stress associated endoplasmic reticulum protein 1 (SERP1). (A) Flow chart. (B) Heat map of differentially expressed mRNAs in pancreatic ductal adenocarcinoma (PDAC) from GEO profiles database. The 10 mRNAs with the most significant difference are shown.

Figure 2

Expression level of the stress associated endoplasmic reticulum protein 1 (SERP1) in pancreatic ductal adenocarcinoma (PDAC) tissues is upregulated. (A) SERP1 was upregulated in six pairs of PDAC tissues compared with adjacent normal tissues. (B) The expression of SERP1 in eight pairs of PDAC tissues was upregulated compared with adjacent normal tissues from GEO database. (C and D) The Human Protein Atlas database results revealed that the expression level of SERP1 were upregulated in PDAC tissues compared with normal tissues. (E) Amplification of SERP1 was found in various tumors, including PDAC.

Figure 3

Correlation between expression level of the stress associated endoplasmic reticulum protein 1 (SERP1) and progression of pancreatic ductal adenocarcinoma (PDAC) patients. (A and B) The expression level of SERP1 in T3/4 stage PDAC specimen was significantly higher than in T1/2 stage, while there was no significant difference between SERP1 and N stage. (C) Correlation analysis was performed between expression level of SERP1 and clinical stages. SERP1 was correspondingly increased with the increase of clinical AJCC stage of PDAC. (D) Constituent ratio with high level of SERP1 was drastically different between normal samples and I/II or III/IV stage of PDAC patients.

Figure 4

The correlation between expression level of the stress associated endoplasmic reticulum protein 1 (SERP1) and prognosis of pancreatic ductal adenocarcinoma (PDAC) patients. (A) K-M curve for OS of patients with high (n=89) and low (n=87) SERP1 expression level. Results revealed that high SERP1 expression level was correlated with shorter OS for PDAC patients. (B) SERP1 high expression group (n=61) had shorter DFS than SERP1 low expression group (n=76). (C) The expression of SERP1 in OS-good and poor patients. (D) The expression of SERP1 in DFS-good and poor patients. (E and F) Multivariate Cox regression analyses showed SERP1 expression was an independent factor of patients' overall survival and disease-free survival.

Figure 5

Stress associated endoplasmic reticulum protein 1 (SERP1) is mainly involved in regulating cell apoptosis. (A) Apoptosis gene sets were enriched in tumor group. The curve is the enrichment score of these genes by GSEA. (B) PANC-1 cells were transfected with three SERP1 siRNA. Forty-eight hours after transfection, the expression level of SERP1 was detected by western blotting, and results revealed that the expression level of SERP1 was obviously downregulated in PANC-1 cells transfected with SERP1-siRNA compared to negative control cells. (C and D) Compared to control group, PANC-1 cells transfected with SERP1-siRNA had a higher rate of apoptosis.

Figure 6

Protein-protein interaction (PPI) and GO analyses for interaction genes with the stress associated endoplasmic reticulum protein 1 (SERP1). (A and B) PPI analysis of interaction genes with SERP1. SRP receptor β subunit (SRPRB) was the main node protein for the interaction network. (C) GO analysis of the interaction gene with SERP1, and these genes were mainly involved in regulating cell death and apoptosis.

Figure 7

Downregulated stress associated endoplasmic reticulum protein 1 (SERP1) promotes apoptosis via increasing SRP receptor β subunit (SRPRB) expression. (A and B) The Human Protein Atlas database results revealed that the expression level of SRPRB were upregulated in stromal fibroblast compared with tumor tissues. (C) Western blot analysis confirmed that downregulated SERP1 can increase the expression of SRPRB in PANC-1 cells. (D) PANC-1 cells were transfected with SRPRB overexpression plasmid, and western blot analysis found that the expression level of SRPRB was significantly increased in PANC-1 cells. (E and F) Compared to control group, SRPRB overexpression significantly promoted apoptosis of PANC-1 cells.

Figure 8

Downregulated stress associated endoplasmic reticulum protein 1 (SERP1) promotes cell apoptosis via regulating SRP receptor β subunit (SRPRB) associated NF-κB activation. (A) NF-κB signaling pathway gene sets were enriched in the tumor group. (B) Correlation analysis revealed that the expression level of SERP1 positively correlated with NF-κB expression in 178 pancreatic ductal adenocarcinoma (PDAC) patients from TCGA database. (C) The expression level of SRPRB was negatively correlated with NF-κB expression in 178 PDAC patients from TCGA database. (D) Western blotting confirmed that downregulated SERP1 could suppress the expression of NF-κB and the phosphorylation NF-κB in PANC-1 cells.