Abstract: PO0895
Integrated Single Nucleus RNA and ATAC-Seq Implicate Cis-Regulatory Chromatin Interactions That Promote Gluconeogenesis in the Human Diabetic Proximal Tubule
Session Information
- Diabetic Kidney Disease: Basic Mechanisms
October 22, 2020 | Location: On-Demand
Abstract Time: 10:00 AM - 12:00 PM
Category: Diabetic Kidney Disease
- 601 Diabetic Kidney Disease: Basic
Authors
- Wilson, Parker C., Division of Anatomic and Molecular Pathology, Department of Pathology and Immunology, Washington University in Saint Louis School of Medicine, Saint Louis, MO, USA, Saint Louis, Missouri, United States
- Muto, Yoshiharu, Division of Nephrology, Department of Medicine, Washington University in Saint Louis School of Medicine, Saint Louis, MO, USA, Saint Louis, Missouri, United States
- Waikar, Sushrut S., Division of Nephrology, Department of Medicine, Boston University School of Medicine, Boston, MA, USA, Boston, Massachusetts, United States
- Humphreys, Benjamin D., Division of Nephrology, Department of Medicine, Washington University in Saint Louis School of Medicine, Saint Louis, MO, USA, Saint Louis, Missouri, United States
Background
Type 2 diabetes is characterized by impaired glucose metabolism, but relatively little is known about cell-specific changes in the kidney. We hypothesized that single nucleus ATAC (snATAC) and RNA (snRNA) sequencing of kidney tissue from patients with early diabetes would reveal cis-regulatory chromatin interactions that promote expression of genes that lead to glucose intolerance.
Methods
We analyzed five kidney samples from patients with early diabetes and five healthy controls. Diabetic patients had elevated A1c and two of five had proteinuria. Serum creatinine (mean = 1.01 mg/dl) was not different between groups. Nuclear preparations were processed with 10x Genomics 5’ v2 or Single Cell ATAC kits, sequenced and counted with Cell Ranger. Analysis was performed with Seurat. snATAC peaks were called with MACS2 using SnapATAC. Chromatin interactions were predicted with Cicero.
Results
A total of 80,576 nuclei were analyzed by snATAC (n=46,564) or snRNA (n=34,012) sequencing and included all cell types. In the diabetic proximal tubule, we observed increased expression of gluconeogenic genes PCK1, ALDOB, FBP1, and G6PC and the sodium bicarbonate exchanger, SLC4A4 (Figure 1; green=upregulated, red=downregulated). Increased expression of GLS and GLUD1 implicate glutamine as a gluconeogenic substrate. Transcriptional changes were accompanied by cell-specific differential chromatin accessibility in regulatory regions that were linked to their respective promoters via predicted chromatin interactions. Differentially accessible regions in the proximal tubule were enriched for NFkB binding motifs, suggesting it may regulate chromatin accessibility in diabetes.
Conclusion
This is the first single cell multi-omic analysis of early human diabetic kidney injury. Our analysis reveals that early diabetes induces changes in chromatin accessibility that promote gluconeogenesis and ammoniagenesis in the proximal tubule, and suggests utility for single cell multi-omic analyses.