ERH Interacts With EIF2 alpha and Regulates the EIF2 alpha/ATF4/CHOP Pathway in Bladder Cancer Cells
FRONTIERS IN ONCOLOGY(2022)
Abstract
Background: There is a lack of research on the molecular interaction of the enhancers of rudimentary homolog (ERH) in bladder cancer (BC) cells. This study aimed to determine the interacting proteins of ERH in human T24 cells. Methods: First, the ERH gene was overexpressed in human T24 cells. Coimmunoprecipitation (co-IP) and shotgun mass spectrometry (MS) analyses were performed to obtain a list of proteins that interact with ERH. Subsequently, bioinformatic analyses with Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and protein-protein interaction (PPI) studies were performed to analyze the ERH-interactive protein list (ERH-IPL). Then, we selected one of the interacting proteins, EIF2 alpha for verification. An immunofluorescence colocalization assay was performed to validate the co-expression of the selected protein, and the binding sites of the two proteins were predicted by ZDOCK technology. Finally, PCR analysis on the downstream molecules of the interacting protein was performed for verification. Results: ERH protein was successfully overexpressed in human T24 cells. We obtained a list of 205 proteins that might directly or indirectly interact with the ERH protein by mass spectrometric analysis. The bioinformatic analysis showed that ERH-interacting proteins were related to "ribonucleoprotein complex ", "ATPase activity ", "nuclear speck ", and "translation factor activity, RNA binding ". We further identified one of the key genes, EIF2S1, and confirmed that the corresponding protein EIF2 alpha is co-expressed and may bind with ERH in human T24 cells. The mRNA levels of molecules ATF4 and CHOP were found to be upregulated by ERH. Conclusion: ERH protein affects "ribonucleoprotein complex ", "ATPase activity ", "nuclear speck ", and "translation factor activity, RNA binding ". The ERH protein can interact with EIF2 alpha and regulate the EIF2 alpha-ATF4/CHOP signaling pathway in human T24 cells.
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Key words
ERH protein, bladder cancer (BC), protein-protein interaction, EIF2 alpha, EIF2a-ATF4/CHOP pathway
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