# Regulation of Cellular Cholesterol Homeostasis

> **NIH NIH R01** · JOHNS HOPKINS UNIVERSITY · 2020 · $473,832

## Abstract

PROJECT SUMMARY
Lipid homeostasis is essential for cell function and disruptions to lipid homeostasis cause disease. Elevated
serum cholesterol is a primary risk factor for heart disease, a leading killer of adults in the United States.
Hepatic fatty acid and triglyceride accumulation promote fatty liver disease that progresses to non-alcoholic
steatohepatitis, liver cirrhosis and cancer. Type II diabetes mellitus is a major risk factor for developing fatty
liver, and alarmingly diabetes is projected to affect one-quarter of the U.S. population by 2050. Understanding
regulation of cellular lipid homeostasis will identify therapeutic opportunities for these common diseases.
 Membrane-bound, basic helix-loop-helix leucine zipper transcription factors called sterol regulatory
element-binding proteins (SREBPs) are the central regulators of cellular lipid homeostasis, controlling
synthesis and uptake of cholesterol, fatty acids, and triglycerides. In Years 1-15 of this project, we successfully
leveraged fission yeast as a simple genetic model for studies of SREBP regulation. We discovered that fungal
SREBP is a conserved oxygen-responsive transcription factor required for adaptation to low oxygen and
virulence of pathogenic fungi. Our work defined two new paradigms for cellular oxygen sensing: (1) oxygen
supply controls sterol synthesis, and (2) oxygen regulates binding of the prolyl hydroxylase Ofd1 to effectors. In
a pathway distinct from mammals, activation of fungal SREBP requires ubiquitination by the Golgi Dsc E3
ligase and cleavage by the rhomboid intramembrane protease Rbd2. A second, unidentified protease is
required to complete release of the N-terminal SREBP transcription factor from the membrane.
 The advent of CRISPR/Cas9 technology enables genetic experiments directly in human cells. Accordingly,
the current proposal reflects a transition in our studies of the SREBP pathway from yeast to mammalian cells.
In Years 16-20, we will (1) complete our description of the yeast SREBP pathway by identifying the second
SREBP protease, (2) test whether oxygen also regulates mammalian SREBP, and (3) deploy CRISPR/Cas9
genetics to identify new regulators of SREBP and lipid homeostasis. We propose the following specific aims:
AIM 1. TO IDENTIFY THE SECOND FISSION YEAST SREBP PROTEASE.
AIM 2. TO TEST WHETHER THE HIF-INSIG2 AXIS REGULATES SREBP IN VITRO AND IN VIVO.
AIM 3. TO IDENTIFY NEW REGULATORS OF SREBP2-N USING CRISPR/CAS9 GENETIC SELECTIONS.
The impact of the proposed studies is high as we will identify a new enzymatic target for antifungal drug
development, define a pathway for hypoxic regulation of mammalian SREBP, and discover novel regulators of
LDL receptor expression. Our team has extensive expertise in studies of hypoxia in yeast and mice, and we
take innovative approaches in applying our knowledge from yeast to mammalian cells. Given the central role
for SREBP in control of lipid homeostasis, our findings will inform both cardiovascular an...

## Key facts

- **NIH application ID:** 9840494
- **Project number:** 5R01HL077588-16
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** PETER J. ESPENSHADE
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $473,832
- **Award type:** 5
- **Project period:** 2004-07-01 → 2022-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9840494

## Citation

> US National Institutes of Health, RePORTER application 9840494, Regulation of Cellular Cholesterol Homeostasis (5R01HL077588-16). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9840494. Licensed CC0.

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