Cbl-b E3 ligase-mediated neddylation and activation of PARP-1 induces vascular calcification

Abstract Aims: Vascular calcification (VC) refers to the accumulation of mineral deposits on the walls of arteries and veins, and it is closely associated with increased mortality in cardiovascular disease, particularly among high-risk patients with diabetes and chronic kidney diseases (CKD). Neuronal precursor cell-expressed developmentally downregulated protein 8 (NEDD8) is an ubiquitin-like protein that plays a pivotal role in various cellular functions, primarily through its conjugation to target proteins and subsequent relay of biological signals. However, the role of NEDDylation in VC has not been investigated. Methods and Results: In our study, we observed that MLN4924, an inhibitor of the NEDD8-activating E1 enzyme, effectively impedes progress of VC. By LC-MS/MS analysis, we identified that poly(ADP-ribose) polymerase 1 (PARP-1) is subjected to NEDD8 conjugation, leading to an increase in PARP-1 activity during VC. Subsequently, we uncovered that the PARP-1 NEDDylation is mediated by the E3 ligase Cbl proto-oncogene B (Cbl-b) and is reversed by the NEDD8-specific protease 1 (NEDP-1) during VC. Furthermore, Cbl-b C373 peptide effectively mitigates the inactive form of E3 ligase activity of Cbl-b, ultimately preventing VC. Conclusions: These findings provide compelling evidence that the NEDD8-dependent activation of PARP-1 represents a novel mechanism underlying vascular calcification and suggests a promising new therapeutic target for VC.