Kidney Week

Abstract: SA-PO597

Possible Link Between Aging Nephropathy and DHTKD1 in C57BL/6J Mice

Session Information

• Noncystic Mendelian Diseases
  November 04, 2017 | Location: Hall H, Morial Convention Center
  Abstract Time: 10:00 AM - 10:00 AM

Category: Genetic Diseases of the Kidney

• 802 Non-Cystic Mendelian Diseases

Authors

• Susnik, Nathan D., Hannover Medical School, Hannover, Germany

Nathan D. Susnik,

• Korstanje, Ron, The Jackson Laboratory, Bar Harbor, Maine, United States

Ron Korstanje,

• Reinholdt, Laura, The Jackson Laboratory, Bar Harbor, Maine, United States

Laura Reinholdt,

• Hanke, Nils, Hannover Medical School, Hannover, Germany

Nils Hanke,

• Hegermann, Jan, Hannover Medical School, Hannover, Germany

Jan Hegermann,

• Wrede, Christoph, Hannover Medical School, Hannover, Germany

Christoph Wrede,

• Bähre, Heike, Hannover Medical School, Hannover, Germany

Heike Bähre,

• Haller, Hermann G., Hannover Medical School, Hannover, Germany

Hermann G. Haller,

• Schiffer, Mario, Hannover Medical School, Hannover, Germany

Mario Schiffer,

• Schmitt, Roland, Hannover Medical School, Hannover, Germany

Roland Schmitt,

Background

During renal aging experiments, we found a spontaneous phenotype in a cohort of 18-22 month old C57BL/6J mice. Old mice developed pronounced glomerulosclerosis, amyloidosis, and renal dysfunction with proteinuria (GARD). Here, we tested the assumption that the GARD phenotype was due to a spontaneous mutation.

Methods

Whole exome sequencing data from a GARD mouse was compared to publicly available C57BL/6J reference sequence (GRCm38). C57BL/6J and C57BL/6JRJ mice purchased from different suppliers were sequenced for mutations in DHTKD1. Cellular localization of DHTKD1 was examined in murine and human kidneys and in renal cells. DHTKD1 expression was manipulated with Crispr-Cas9 and siRNA in cell lines and morpholino in zebrafish.

Results

Whole exome sequencing revealed that GARD mice had a single nucleotide polymorphism leading to an amino acid substitution (A335T) in the thiamine-binding motif of dehydrogenase E1 and transketolase domain containing 1 (DHTKD1). All C57BL/6J mice in the GARD cohort were homozygous for the Dhtdk1 mutation (Dhtkd1^A335T/A335T) while C57BL/6JRJ mice with normal renal aging carried the reference allele (Dhtkd1^+/+). DHTKD1 was expressed in podocytes, proximal tubules, and distal tubules. Contrary to previously reported data, knockout (KO) of DHTKD1 left mitochondrial morphology, genes associated with mitochondrial function, and ATP production unchanged. Flow cytometry, however, revealed more MTC02 in DHTKD1 KO cells. KO cells also had more 2-aminoadipic- and 2-oxoadipic acid, metabolites of lysine degradation. siRNA in human podocytes changed some mitochondria-associated genes, but made no difference in ATP production. Knockdown of Dhtkd1 in zebrafish larvae led to proteinuria and podocyte foot process effacement. While these changes indicate a functional role for DHTKD1 in renal maintenance, C57BL/6J Dhtkd1^A335T/A335T mice aged at our own facilities did not develop GARD.

Conclusion

Overall, knockdown of DHTKD1 causes structural and functional changes in cells and zebrafish; however, the GARD phenotype in Dhtkd1^A335T/A335T mice was not fully penetrant, suggesting that other variables like diet or housing are involved.

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Funding

• Other NIH Support