Deficiency of NARFL increases transcription of NADPH oxidases and ROS production impairing the function of endothelial cells

Abstract

AimsNuclear prelamin A recognition factor-like (NARFL) is involved in cytosolic iron‑sulfur (FeS) protein biogenesis and cellular defense against oxidative stress. Previous study reported that increased oxidative stress and subintestinal vessel (SIV) malformation in narfl knockout zebrafish. However, the underlying mechanism of oxidative stress caused by NARFL deficiency remains unclear. The present study was sought to investigate the function of NARFL in endothelial cells. MethodsNARFL knockdown assay was performed in two cell lines and NADPH oxidase (Nox) were measured using Western blotting. Nox inhibitors were selected for assessing the potential sources of reactive oxygen species (ROS) generation. Cell migration was detected using wound healing assay and transwell assay. Cell cycle was analyzed using flow cytometry. Promoter activity assay and Chromatin immunoprecipitation (ChIP) assay were chosen for investigating the molecular mechanism of Nox transcription. ResultsNARFL deficiency resulted in upregulated expressions of Nox2, Nox4, and p47phox and increased ROS levels in endothelial cells. Nox2 knockdown reversed the effects and improved endothelial dysfunctions caused by NARFL deficiency. ChIP experiments revealed that NARFL knockdown increased the recruitment of RNA polymerase II and modification of histones at the promoter sites of Nox2 and Nox4. ConclusionNARFL knockdown induced the transcriptional activation of Nox2 and Nox4, which resulted in increased ROS levels and impaired endothelial functions.