Kidney Week

Abstract: FR-PO353

Proximal Tubule-Specific KLF6-Mediated Amino Acid Metabolism Is Critical for the Progression of Kidney Injury

Session Information

• CKD: Mechanisms - II
  November 08, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: CKD (Non-Dialysis)

• 2103 CKD (Non-Dialysis): Mechanisms

Authors

• Piret, Sian, Stony Brook Medicine, Stony Brook, New York, United States

Sian Piret,

• Guo, Yiqing, Stony Brook Medicine, Stony Brook, New York, United States

Yiqing Guo,

• Mallipattu, Sandeep K., Stony Brook Medicine, Stony Brook, New York, United States

Sandeep K. Mallipattu,

Background

Transcriptional regulators of DNA-damage pathways leading to renal fibrosis are not well characterized. Krüppel-like factor 6 (KLF6), a zinc finger transcription factor, is expressed in the proximal tubule (PT) and highly upregulated in renal ischemia-reperfusion injury, sepsis, and DNA damage models, but its role in PT injury and tubulointerstitial fibrosis is unknown.

Methods

PT-specific Klf6 knockdown (Klf6^PTKO) mice were generated by breeding Klf6^fl/fl and Pepck-Cre mice (controls- Klf6^fl/fl littermates). The “Tet-On” system was used to generate global inducible human KLF6 (hKLF6) overexpressing mice (controls- TRE-hKLF6). Mice were given AAI or vehicle DMSO i.p. every 3 days for 2-3 weeks and euthanized 3 days after the final injection for functional, histological, RNA-Seq and ChIP-enrichment analysis of renal cortex.

Results

AAI-treated Klf6^PTKO mice had lower serum creatinine and urea nitrogen, preserved PT, and reduced inflammation and fibrosis, versus AAI-treated Klf6^fl/fl mice. Conversely, AAI-treated hKLF6 mice had worse renal function, more loss of PT, and increased inflammation and fibrosis versus AAI-treated control mice. RNA-Seq analysis identified 388 genes that were downregulated in Klf6^fl/fl mice and significantly higher (preserved) in Klf6^PTKO mice after AAI. These were predominantly metabolic genes, including in amino acid metabolic and fatty acid oxidative pathways. ChIP-enrichment analysis of genes with KLF6 binding sites <1kb from a transcription start site (TSS) enriched for genes encoding enzymes critical to the essential branched-chain amino acid (BCAA) pathways, suggesting direct regulation by KLF6. AAI inhibits mitochondrial function causing production of reactive oxygen species, and the BCAA pathways preserved in Klf6^PTKO mice supply TCA cycle intermediates (e.g. succinyl-CoA), and enhance glutathione production, which may counteract the effects of AAI. Analysis of expression arrays of human CKD biopsies showed similar downregulation of BCAA genes with KLF6 binding sites close to the TSS, versus control samples.

Conclusion

Loss of PT KLF6 was associated with preserved BCAA metabolic enzymes that may help maintain the TCA cycle and increase glutathione, thus reducing injury after AAI. The potential role of transcriptional regulation of amino acid metabolism in driving PT injury has not previously been described.

Funding

• NIDDK Support