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  • Silencing of ER-resident oxidoreductase PDIA3 inhibits malignant biological behaviors of multidrug-resistant gastric cancer.

Silencing of ER-resident oxidoreductase PDIA3 inhibits malignant biological behaviors of multidrug-resistant gastric cancer.

Acta biochimica et biophysica Sinica (2021-08-08)
Danyang Song, Meng Guo, Kaichu Wu, Jianyu Hao, Yongzhan Nie, Daiming Fan
ABSTRACT

Glycosylation is a common posttranslational modification of proteins, which plays a role in the malignant transformation, growth, progression, chemoresistance, and immune response of tumors. Disulfide isomerase family A3 (PDIA3) specifically acts on newly synthesized glycoproteins to promote the correct folding of sugar chains. Studies have shown that PDIA3 participates in multidrug-resistant gastric cancer (MDR-GC). In this study, we performed western blot analysis and immunohistochemistry to identify PDIA3 expression. Cell proliferation was assessed by CCK-8 assay. Transwell assays were used to detect the migration and invasion abilities of cells. Immunoprecipitation coupled to mass spectrometry (IP-MS) analysis was employed to identify PDIA3-interacting proteins and the associated pathways in MDR-GC cells. Glycoprotein interactions and translocation were detected by immunofluorescence assay. The results showed that PDIA3 knockdown significantly inhibited the proliferation, invasion, and migration abilities of MDR-GC cells. Kyoto Encyclopedia of Genes and Genomes analysis of the IP-MS results showed that PDIA3 was closely associated with focal adhesion pathways in MDR-GC cells. Additionally, important components of focal adhesion pathways, including fibronectin-1 (FN1) and integrin α5 (ITGA5), were identified as pivotal PDIA3-binding glycoproteins. Knockdown of PDIA3 altered the cellular locations of FN1 and ITGA5, leading to abnormal accumulation. In conclusion, our results suggest that knockdown of PDIA3 inhibited the malignant behaviors of MDR-GC cells and influenced the translocation of FN1 and ITGA5.

MATERIALS
Product Number
Brand
Product Description

Millipore
Protease Inhibitor Cocktail Set III, EDTA-Free, Protease inhibitor cocktail III, EDTA-free for inhibiting aspartic, cysteine, and serine proteases as well as aminopeptidases in mammalian cells and tissues.
Sigma-Aldrich
Anti-β-Actin antibody, Mouse monoclonal, clone AC-15, purified from hybridoma cell culture
Millipore
Phosphatase Inhibitor Cocktail Set II, A cocktail of five phosphatase inhibitors for the inhibition of acid and alkaline phosphatases as well as protein tyrosine phosphatases (PTPs). Suitable for use with cell lysates and tissue extracts.