c-Ski is a novel repressor of NF-κB through interaction with p65 and HDAC1 in U937 cells.

The nuclear factor kappa B (NF-κB) signalling pathway plays a crucial role in the regulation of inflammation, and previous research from our lab and others suggests that c-Ski has potential anti-inflammatory effects. However, the role and mechanism of c-Ski, which are related to the regulation of the NF-κB pathway, are still unclear. Here, U937 cells were used, and increasing c-Ski protein levels inhibited inflammatory factor production, invasion, and phagocytosis. The anti-inflammatory effect of c-Ski was similar to that of hormones. Subsequently, immunoprecipitation (IP), Western blot (WB), electrophoretic mobility shift assays (EMSAs), and dual-luciferase reporter assays were used to determine whether increasing c-Ski protein levels could increase c-Ski binding to NF-κB p65 (p65), leading to a decrease in the acetylation level and transcriptional activity of p65. Conversely, decreased p65 expression through targeted small interfering RNA (siRNA) caused the loss of the anti-inflammatory effects of c-Ski. Furthermore, immunoprecipitation confirmed the mutual interaction of c-Ski with HDAC1 and p65, and WB revealed that the anti-inflammatory effect of c-Ski was achieved through the deacetylation of p65 by HDAC1 combined with HDAC1 siRNA and inhibitors. Additionally, through quantitative proteomic analysis, we determined that increasing c-Ski levels had inhibitory effects on the NF-κB pathway. Finally, similar results were also obtained using primary bone marrow-derived macrophages (BMDMs). These findings not only confirm the anti-inflammatory effect of c-Ski but also reveal novel molecular pathways and regulatory molecules of c-Ski, which may be promising targets for direct intervention in the inflammatory response through regulation of c-Ski.