Abstract: PO1594
The Role of Claudin Variants in the Formation of Kidney Stones
Session Information
- Genetic Diseases of the Kidneys: Non-Cystic - 1
October 22, 2020 | Location: On-Demand
Abstract Time: 10:00 AM - 12:00 PM
Category: Genetic Diseases of the Kidneys
- 1002 Genetic Diseases of the Kidneys: Non-Cystic
Authors
- Zhuang, Yuan, McGill University Faculty of Medicine, Montreal, Quebec, Canada
- El andalousi, Jasmine, Research institute of McGill university, Montreal, Quebec, Canada
- Andonian, Sero, McGill University Faculty of Medicine, Montreal, Quebec, Canada
- Jean-Claude, Bertrand Jacques, McGill University Faculty of Medicine, Montreal, Quebec, Canada
- Ryan, Aimee K., McGill University Faculty of Medicine, Montreal, Quebec, Canada
- Gupta, Indra R., McGill University Faculty of Medicine, Montreal, Quebec, Canada
Background
Genetic risk factors contribute to the formation of calcium-based kidney stones. The majority of calcium is reabsorbed via paracellular transport through tight junctions along the human nephron epithelium where Claudin proteins are expressed. Claudins determine the selectivity and permeability of different nephron segments. Studies have shown that CLDN gene sequence variants are associated with kidney stones. I hypothesize that sequence variants in Claudin genes that regulate paracellular renal transport of calcium. will be associated with the formation of kidney stones.
Methods
Patient DNA was analyzed by Fluidigm Next Generation Sequencing and confirmed by sanger sequencing. Rare variants (MAF<1%) were compared to the gnomAD database. In silico prediction software was used to predict the impact of the amino acid change. Human claudin variants were generated by site-directed mutagenesis and cloned into a mammalian expression vector, pEGFP. Immunofluorescence was performed on HEK293 cells that were transiently transfected with both variant and WT sequences.
Results
Ninety adult patients (45 females, 45 males) with recurrent calcium-based kidney stones were recruited from one urologist’s kidney stone clinic. Seventy-two percent (65/90) of the patients self-defined as Canadian-European. Sixty-two percent (56/90) of the patients presented with the first kidney stone less than 40 years of age. Four novel heterozygous missense variants were identified in the following: CLDN11 S157F, CLDN16 K29E, CLDN17 A94V, and CLDN18 H212D. Nine rare variants include CLDN4 A82T, CLDN4 A113T, CLDN7 V55I, CLDN8 A94V, CLDN8 M97T, CLDN12 M98V, CLDN23 A90T, and CLDN24 V97I. CLDN4 A82T, CLDN8 A94V, CLDN11 S157F, and CLDN17 A94V are predicted to be deleterious.
HEK293 cells were transiently transfected with CLDN4 A82T and the mutant protein was unable to localize to the tight junction, unlike the WT CLDN4 protein which did co-localize with ZO-1 by immunofluorescence as expected (n=3 independent experiments). Other claudin variants are under evaluation.
Conclusion
The rare heterozygous variant, CLDN4 A82T is located at the second transmembrane domain and predicted to be deleterious. Functional analysis showed that CLDN4 A82T has an impact on the localization of the protein to the tight junction.
Funding
- Private Foundation Support