Cellular lipid homeostasis is required to maintain bilayer fluidity, membrane impermeability, and organelle identity. Disturbances in systemic lipid homeostasis lie at the core of the pathologies for both coronary artery disease and obesity related type II diabetes. The long-term goal of this research is to translate knowledge of cellular regulatory events to that of the whole organism and to advance our understanding of these increasingly widespread diseases. As a first step toward this goal, we will use cholesterol as a model lipid to understand how cells measure levels of these largely insoluble molecules and in turn modulate their production. Cholesterol homeostasis in mammalian cells is regulated by a feedback mechanism that monitors the level of cholesterol in membranes and alters transcription of genes required for cholesterol supply. Transcription of these genes is controlled by the ER membrane-bound transcription factor called SREBP that is activated and released from the membrane by proteolysis in sterol-depleted cells. To accelerate discovery of sterol homeostasis regulators, we are studying the SREBP pathway in the fission yeast Schizosaccharomyces pombe. Yeast SREBP, called Sre1, functions in a new oxygen sensing pathway that mediates adaptation of cells to low oxygen. Interestingly, sterols regulate Sre1 activity through a novel mechanism in fission yeast, and evidence also indicates that Sre1 cleavage is mediated by a unique proteolytic system. In this project, a combination of genetic, molecular, and biochemical approaches will be used to accomplish the following specific aims: 1) To identify genes required for Sre1 cleavage using a genetic selection;2) To define the machinery for Sre1 cleavage;and 3) To define the mechanism of sterol-regulated Sre1 cleavage. The long-term goal of this project is to use S. pombe as a genetic model to understand how cells measure levels of insoluble, membrane-embedded cholesterol. The expectation is that these studies will describe new mechanisms for lipid sensing and proteolysis that will advance our understanding of the mammalia SREBP pathway and eukaryotic cell biology.

Public Health Relevance

Heart disease is a leading killer of adults in the United States. Proper regulation of cellular cholesterol homeostasis is integral to cardiovascular health. In this project, we will use yeast as a genetic model organism to define new mechanisms for regulation of cholesterol homeostasis with the goal of identifying improved therapeutic strategies for lowering serum cholesterol and preventing heart disease.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL077588-09
Application #
8253717
Study Section
Membrane Biology and Protein Processing (MBPP)
Program Officer
Liu, Lijuan
Project Start
2004-07-01
Project End
2014-04-30
Budget Start
2012-05-01
Budget End
2013-04-30
Support Year
9
Fiscal Year
2012
Total Cost
$413,009
Indirect Cost
$161,174
Name
Johns Hopkins University
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
001910777
City
Baltimore
State
MD
Country
United States
Zip Code
21218
Tong, Zongtian; Kim, Min-Sik; Pandey, Akhilesh et al. (2014) Identification of candidate substrates for the Golgi Tul1 E3 ligase using quantitative diGly proteomics in yeast. Mol Cell Proteomics 13:2871-82
Shao, Wei; Espenshade, Peter J (2014) Sterol regulatory element-binding protein (SREBP) cleavage regulates Golgi-to-endoplasmic reticulum recycling of SREBP cleavage-activating protein (SCAP). J Biol Chem 289:7547-57
Brookheart, Rita T; Lee, Chih-Yung S; Espenshade, Peter J (2014) Casein kinase 1 regulates sterol regulatory element-binding protein (SREBP) to control sterol homeostasis. J Biol Chem 289:2725-35
Lloyd, S Julie-Ann; Raychaudhuri, Sumana; Espenshade, Peter J (2013) Subunit architecture of the Golgi Dsc E3 ligase required for sterol regulatory element-binding protein (SREBP) cleavage in fission yeast. J Biol Chem 288:21043-54
Porter, Joshua R; Burg, John S; Espenshade, Peter J et al. (2012) Identifying a static nonlinear structure in a biological system using noisy, sparse data. J Theor Biol 300:232-41
Stewart, Emerson V; Lloyd, S Julie-Ann; Burg, John S et al. (2012) Yeast sterol regulatory element-binding protein (SREBP) cleavage requires Cdc48 and Dsc5, a ubiquitin regulatory X domain-containing subunit of the Golgi Dsc E3 ligase. J Biol Chem 287:672-81
Yeh, Tzu-Lan; Lee, Chih-Yung S; Amzel, L Mario et al. (2011) The hypoxic regulator of sterol synthesis nro1 is a nuclear import adaptor. Structure 19:503-14
Burg, John S; Espenshade, Peter J (2011) Regulation of HMG-CoA reductase in mammals and yeast. Prog Lipid Res 50:403-10
Lee, Chih-Yung S; Yeh, Tzu-Lan; Hughes, Bridget T et al. (2011) Regulation of the Sre1 hypoxic transcription factor by oxygen-dependent control of DNA binding. Mol Cell 44:225-34
Burg, John S; Espenshade, Peter J (2011) Glucose controls phosphoregulation of hydroxymethylglutaryl coenzyme A reductase through the protein phosphatase 2A-related phosphatase protein, Ppe1, and Insig in fission yeast. J Biol Chem 286:27139-46

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