Project Details
Disease mechanisms of podocyte injury caused by mutations in genes encoding proteins of the tRNA modifying KEOPS complex.
Applicant
Privatdozentin Dr. Daniela Anne Braun
Subject Area
Nephrology
Term
from 2017 to 2021
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 391152220
Chronic kidney disease (CKD), affecting approximately 10% of the world population, is a major global public health burden. In a subset of CKD, damage to the renal glomerular filter causes substantial loss of plasma proteins in the urine, thus resulting in hypoalbuminemia, generalized edema, and the clinical presentation of nephrotic syndrome. If refractory to treatment, the disease inevitably progresses to end-stage renal failure requiring renal replacement therapy or transplantation for survival. The identification of human genes that, if mutated, cause monogenic forms of nephrotic syndrome has provided novel insights into its pathogenesis and has identified glomerular podocytes, specialized epithelial cells, as the primary site of damage. Podocytes are terminally differentiated cells with very limited regenerative capacity. Consequently, injured podocytes cannot be replenished and loss of more than 20% of podocytes causes irreversible glomerulosclerosis. In genetic studies, we identified mutations in LAGE3, OSGEP, TP53RK, or TPRKB encoding the 4 subunits of the evolutionarily highly conserved KEOPS complex as novel monogenic causes of steroid-resistant nephrotic syndrome with microcephaly in 31 unrelated families. The KEOPS complex mediates an essential posttranscriptional modification of tRNA, known as t6A modification that crucial for accuracy and efficiency of protein translation at the ribosome. Our preliminary data show that knockdown of different genes of the KEOPS complex causes ER stress and apoptosis in immortalized human podocytes. We hypothesize that these two mechanisms contribute to the pathogenesis of KEOPS-related nephrotic syndrome. Our first objective is to perform an in-depth analysis of the molecular mechanisms of podocyte injury following knockdown of different KEOPS genes. In particular, we will analyze the impact of KEOPS-related ER stress on other cross-talking signaling pathways with high relevance for podocyte function as well as on posttranslational processing of podocyte slit-diaphragm proteins. Furthermore, we will study how error-prone protein translation in these cells affects the cellular energy balance and whether, as seen in other diseases, misfolded proteins accumulate. In a next step, we will test different pharmacological strategies for their efficacy in podocytes with KEOPS gene knockdown and establish a fluorescent ER stress reporter podocyte cell line for high-throughput drug-screening. In order to determine the function of the KEOPS complex in mature, terminally differentiated podocytes and during glomerular development, we will generate a podocyte-specific knockout mouse for the gene Osgep. Using these mice, we will determine relevant pathophysiological features of KEOPS-related glomerular disease, perform proteomics to analyze the impact of KEOPS complex dysfunction on the podocyte proteome at different ages, and test therapeutic interventions in vivo.
DFG Programme
Research Grants