Immunoprecipitation-mass Spectrometry Reveals New Functions of RCC2 in Breast Cancer Cell Line MDA-MB-468

Objective To analyze the protein interactome and reveal the functional regulation network of RCC2 protein in MDA-MB-468 cell line.

Methods Using enzyme digestion combined with one generation sequencing, we successfully constructed overexpression vector of RCC2 protein with a Flag tag. MDA-MB-468 cell line with Flag-RCC2 stably overexpression was constructed using a lentiviral vector system. Flag-tag gel beads affinity precipitation was used to collect RCC2-interacting proteins for high-sensitivity mass spectrometry analysis. And bioinformatic analyses of interacting-proteins, including GO annotation, KEGG pathway enrichment, construction of protein-protein interaction network, were further processed.

Results Clustering analyses of 164 proteins with high confidence indicated that RCC2 and its interacting proteins were significantly enriched in ribosomal-related pathways and had the abilities to bind with RNA and ribosomal proteins, revealing its involvement in protein translation and maturation.

Conclusion We systematically identified the RCC2 interactome map and revealed that RCC2 plays a new role of translational regulation in breast cancer cells.