Liquid–Liquid Phase Separation-Related STAT1 Modulating Ubiquitination Aggravates the Progression of Diabetic Nephropathy via Promoting Inflammatory Response

This study aims to screen liquid-liquid phase separation (LLPS)-related genes modulating ubiquitination for diagnosing patients with diabetic nephropathy (DN). After performing differential gene expression analysis, ssGSEA, and immune infiltration analysis, LLPS-related genes modulating ubiquitination, key immune cells, and immune-related pathways were acquired. The correlations between LLPS-related genes and ubiquitination, key immune-related pathways were explored, respectively. The diagnostic genes were identified by four machine learning algorithms, followed by evaluation to predict the performance of the machine learning models on ubiquitination using receiver operating characteristic curves and a confusion matrix. In vitro experiments were applied to explore the function of signal transducer and activator of transcription 1 (STAT1) in DN. Totally, 513 differentially expressed genes were screened between the DN and control groups. The activity of the ubiquitin-mediated proteolysis pathway exhibited a significant difference between the DN and control samples. The fraction of a total of 10 immune cells exhibited a significant difference between the lower and higher ubiquitination groups. Totally, nine LLPS-related genes modulating ubiquitination were acquired, and these genes modulating ubiquitination were enriched in ubiquitin pathways and significantly positively correlated with the complement, IFN-α, and IFN-γ pathways. Two diagnostic genes were obtained, namely dual specificity tyrosine phosphorylation regulated kinase 2 (DYRK2) and STAT1. Notably, downregulation of STAT1 effectively alleviated IFN-γ, IFN-α, IL-1β, and TNF-α levels in D-glucose-treated HK-2 cells, as well as the protein level of FK2. LLPS-related STAT1 modulating ubiquitination accelerates the inflammatory response in DN.