Not Everything Is “Genetic”, but Genes Are Involved in Everything (adapted from Kenneth M. Weiss). Our group is interested in identification and investigation of genetic, clinical, and environmental factors determining onset of chronic kidney disease (CKD) and kidney survival. We make use of next-generation sequencing techniques and deep-phenotyping to identify genetic variants that are predictive for disease progression or convey protection from organ failure. We functionally evaluate identified germline variants in vitro in order to understand underlying molecular mechanisms leading to CKD on the one hand or protecting from kidney failure on the other. By doing so, we aim at defining and targeting molecular switches responsible for health maintenance and disease alleviation.
One of six patients undergoing renal transplantation has autosomal-dominant polycystic kidney disease (ADPKD) caused by heterozygous germline mutations in one of two main disease genes, namely PKD1 (encoding polycystin 1, PC1) or PKD2 (encoding polycystin 2, PC2).ADPKDis the commonest genetic disorder leading to CKD including end-stage kidney failure (ESKF).1-3 ESKF from ADPKD commonly occurs between ages 30-80 years. While PKD1-associated disease is generally more severe than PKD2-disease, exemplified by a 20-year difference in mean age at ESKF (55 versus 75 years), current genotype-phenotype correlations are still crude.4,5 For example, some patients with even identical PKD1-mutations vary dramatically in their progression. We demonstrated that additional non-diagnostic hypomorphic PKD1-germline variants, as well as variants in genes involved in proteostasis mechanistically, add to the PKD1-mutational effect.6,7 The underlying mechanisms involve posttranslational modification and endoplasmic reticular (ER)-processing of PC1. As a result, PC1 expression at the cell surface is reduced.8 Conversely, opposite mechanisms may confer renoprotection by increasing PC1 surface expression. We aim to learn from clinical outliers and hypothesize that the latter mechanisms play a role in individuals with ADPKD not depending on dialysis until their 70s and beyond. To address this, we propose the following two specific aims and work packets (WP):
Aim 1/WP1: Identification of genetic determinants associated with non-progression and kidney survival. Exome sequencing in individuals (n=20) with mildest, non-progressive ADPKD-PKD1 (non-ESKF > 70 yrs) and most severe ADPKD-PKD1 (ESKF < 40 yrs) from the Leipzig-Berlin ADPKD-cohort (n=400) via the BIH-sequencing core facility. Consecutive assessment for additional rare and common variants (single variant analysis and variant burden analysis) by use of a predefined CKD/ADPKD-candidate gene panel. Identification of differentially enriched candidate gene sets and single variants, suitable for functional analysis (WP2).
Aim 2/WP2: Validation of genetic determinants associated with non-progression and kidney survival. Overexpression of two most promising gene variants significantly associated with kidney survival and consecutive use of established cellular read-outs on RNA and protein level. Planned analyses include qRT-PCR, Western Blot, and immunofluorescence imaging (e.g. PC1 surface expression).