Kidney Week

Abstract: TH-OR078

HIF Regulation of Nephron Progenitor Metabolic State Mediates Cell Fate Decisions

Session Information

• Novel Mechanisms and Pathways in Kidney Development
  November 02, 2017 | Location: Room 385, Morial Convention Center
  Abstract Time: 04:30 PM - 04:42 PM

Category: Developmental Biology and Inherited Kidney Diseases

• 401 Developmental Biology

Authors

• Murali, Anjana, University of Pittsburgh, Pittsburgh, Pennsylvania, United States

Anjana Murali



Anjana Murali is a senior undergraduate student at the University of Pittsburgh and a guaranteed admit medical student to Pitt Med. She has been a part of the Sims-Lucas lab at the Children’s Hospital of Pittsburgh for the past 3 years, where she has been interrogating the role of oxygen-dependent signaling during kidney development. Last year, Murali was named a Barry M. Goldwater Honorable Mention winner for her excellence in research and a Harry S. Truman Finalist for her leadership and commitment to public service. She has been a strong advocate for gender equity since founding Queen’s Game, the first all girls’ chess camps in the United States, where she motivates girls to play chess and helps them realize their potential to shatter the glass ceiling. Murali hopes to translate that into the STEM fields and increase the number of females and underrepresented groups in science. Through her work with the Youth Research Advisory Board, she is also working to make scientific research more transparent to the public. Murali aspires to become a physician-scientist who helps influence health policies for women and marginalized communities.

• Cargill, Kasey, University of Pittsburgh, Pittsburgh, Pennsylvania, United States

Kasey Cargill,

• Mukherjee, Elina, University of Pittsburgh, Pittsburgh, Pennsylvania, United States

Elina Mukherjee,

• Saifudeen, Zubaida R., Tulane University School of Medicine, New Orleans, Louisiana, United States

Zubaida R. Saifudeen,

• Sims-Lucas, Sunder, Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania, United States

Sunder Sims-Lucas,

Background

Hypoxia inducible factors (HIFs) are transcription factors involved in the differentiation of nephron progenitors (NP) into functional nephrons. Alterations in nephron differentiation lead to renal abnormalities. As renal oxygen increases, von Hippel-Lindau (VHL) marks HIF-1α for degradation and facilitates normal nephron differentiation. Alternatively, pathological hypoxia results in HIF-1α accumulation and initiation of processes including cellular survival and metabolism. Previous in vitro studies have also linked HIF-1α stabilization to mitochondrial pathologies suggesting that HIF-1α plays a role in cellular and mitochondrial respiration. Therefore, we hypothesize that HIF-1α alters mitochondrial function, mediating a metabolic switch to determine NP fate.

Methods

To determine the role of HIFs in the NPs we utilized VHL floxed mice bred with the Six2EGFPcre line, to generate Six2creVHL^lox/lox mutant mice. RNA sequencing was conducted on E14 whole kidneys to look at differential gene expression, and validated via RT-PCR. We also performed a metabolic assessment using seahorse extracellular flux analysis, and coupled this with immunofluorescence (IF) and western blot analysis to analyze metabolic markers.

Results

RNA sequencing of Six2creVHL^lox/lox mutant mice revealed metabolic gene dysregulation. Furthermore, seahorse extracellular flux analysis suggested that mutants with HIF-1α stabilization in the NPs remained in a glycolytic state, and were subsequently unable to switch to oxidative phosphorylation, which drives NP differentiation. IF staining and western blot analysis supported these results revealing fewer mature nephron structures and decreased mitochondrial content in mutant kidneys.

Conclusion

HIFs are critical in determining the metabolic state of the NPs. In the absence of VHL, HIF1α is stabilized. This stabilization maintains NPs in a state of glycolysis and in turn blocks the switch to oxidative phosphorylation causing NP differentiation defects and kidney malformations.

Funding

• NIDDK Support