Transcription factor EP300 targets SIRT5 to promote autophagy of nucleus pulposus cells and attenuate intervertebral disc degeneration

Abstract

Background

Intervertebral disc degeneration (IVDD) is a prevalent spinal ailment and the leading cause of chronic low back pain. Understanding the exact pathogenesis of IVDD and developing targeted molecular drugs will be important in the future. Autophagy plays a key role in the metabolic processes and in the quality control of proteins in IVDD. However, the role of autophagy in the senescence of nucleus pulposus cell (NPC), the primary cells in the intervertebral disc responsible for maintaining the disc's structure and function, is not yet clear.

Methods

Gene expression profiling data of human disc tissue were obtained from the Gene Expression Omnibus GSE15227, GSE23130, and GSE70362 datasets. Autophagy-related differentially expressed genes were identified from the Molecular Signatures Database (MSigDB) database. Weighted gene co-expression network analysis (WGCNA), receiver operating characteristic (ROC) curves, and least absolute shrinkage and selection operator (LASSO) regression identified an autophagy-related hub gene that encodes the E1A binding protein EP300 transcription factor in IVDD samples. Potential downstream target genes of EP300 were identified by bioinformatics analysis. The analysis identified sirtuin 5 (SIRT5) as a potential downstream target of EP300. Chromatin immunoprecipitation (ChIP)-qPCR, small interfering RNA (siRNA), and luciferase reporter gene assays were used to verify the interaction of EP300 and SIRT5 in vitro. For in vivo experiments, SIRT5 knockout mice and SIRT5-overexpressing adeno-associated virus serotype 5 (AAV5) were constructed to verify the effect of the EP300–SIRT5 signal axis on the progression of IVDD.

Results

EP300 expression was reduced in the IVDD samples compared with its expression in healthy disc tissue samples. The reduced EP300 expression inhibited the occurrence of autophagy, which promoted NPC senescence. ChIP-qPCR and luciferase reporter gene assays showed that EP300 promoted SIRT5 expression by direct binding to its promoter. Activation of EP300 expression increased SIRT5 expression and significantly improved autophagy for inhibition of NPC senescence. In vivo experiments confirmed that knockdown of EP300 promoted NPC senescence and led to an exacerbation of IVDD, which was reversed by SIRT5 overexpression.

Conclusion

Our results provide the first evidence for the importance of EP300 and SIRT5 interactions in promoting IVDD development by inhibiting autophagy during IVDD. The EP300–SIRT5 signaling axis was identified as a promising target for therapy of IVDD based on autophagy genes.