The NFE2L1 transcription factor is a key modulator of the stress response in podocytes

Abstract

Background
Chronic kidney disease (CKD) is a major public health problem, particularly in the elderly. Injury to any part of the nephron cause kidney failure. However, glomerular damage, caused by primary glomerular disease or conditions including diabetes and hypertension, is the most frequent cause of CKD. Podocytes are highly specialised terminally differentiated epithelial cells covering the glomerular basement membrane and are a vital component of the glomerular filtration barrier. It is thought that podocyte damage, injury and eventual loss is the principal mechanism of initiation and progression of CKD. There is relatively little known about the mechanism which drives podocyte stress response, damage, and eventual loss. Nuclear Factor, Erythroid 2 Like 1 (NFE2L1) is a complex multi-isoform transcription factor known as the master regulator of basal expression of antioxidant genes and the proteasome. Herein, we hypothesised that the nuclear expression of the NFE2L1 transcription factor is crucial in modulating the podocyte stress response to injury, and reduced expression may be associated with podocyte damage and subsequent progression of kidney disease.

Methods
We investigated NFE2L1 expression levels in normal human kidney podocytes and biopsies from various kidney disease patients, utilising image analysis of multiplexed immunofluorescence proteins. We then studied NFE2L1 isoform expression in conditionally immortalised podocytes cell lines. Finally, using chromatin immunoprecipitation (ChIP) we aimed to determine NFE2L1 transcriptional targets.

Results
The expression level of the nuclear NFE2L1 isoform is drastically reduced in podocytes in different renal disease biopsies compared to that of normal kidney. Its reduced expression in podocyte nuclei coincides with a reduction in antioxidant NAD(P)H dehydrogenase NQO1 protein. Furthermore, we identified the presence of NFE2L1 transcriptionally active isoforms in differentiated podocyte and that upon treatment with nephrotoxic agent, puromycin aminonucleoside (PAN), led to diminished levels of nuclear NFE2L1. Using chromatin immunoprecipitation (ChIP) and real-time quantitative PCR, we established that NQO1 is a direct transcriptional target of NFE2L1.

Significance
Taken together, we have established that NFE2L1 expression is of vital importance to in maintaining podocyte homeostasis, and its modulation is associated with a change in podocyte-specific markers and podocyte health. Podocyte damage is acknowledged as a critical factor in the pathogenesis of kidney disease and therefore identifying a potential pharmaceutical targets may help to develop agents to treat glomerulopathies.

Date of Award	17 Jun 2022
Original language	English
Awarding Institution	University of St Andrews
Supervisor	David James Harrison (Supervisor) & Paul Andrew Reynolds (Supervisor)