The concentration of intracellular free sterol is strictly regulated to optimize the fluidity and function of all eukaryotic membranes. This homeostasis is imparted by several mechanisms, the crux of which is sensing of sterol and transport of this molecule from sites of synthesis or uptake to intracellular organelles where it can be metabolized or utilized. Aberrations in these processes lead to multiple disease pathologies, including atherosclerosis and Niemann Pick C (NPC) disease. In this project, genetic strategies in yeast will be applied to study sterol transport and sensing as it pertains to human NPC disease. Using a candidate gene approach based on sequence conservation between humans and yeast, we have identified an excellent yeast homolog to the gene defective in Niemann Pick type C disease (NPC1). There are two principle aims to this project.
Aim 1 will define the role of the Niemann-Pick C type 1 gene family in intracellular sterol transport. This will test the hypothesis that NPC1 and its yeast homolog, Ncr1, mediate vesicle fusion events and/or act as sterol responsive signal transducers. Molecular, genetic and biochemical approaches including site-directed mutagenesis and sub-cellular protein localization, will be applied.
Aim 2 has two components that target pathways that interact in NPC disease to mediate sterol transport and sensing. Firstly, to characterize components of sterol transport pathways by virtue of interactions with the NCR1 gene product that effect normal cell viability. The second component will isolate constituents of the NPC/Ncr-complex to test the hypothesis that key steps in sterol transport and its regulation involve protein-protein interactions. Factors that complex with NCR1 and NPC1 will be identified using the yeast two-hybrid system. The NPC2 gene may arise from this screen. These studies will likely elucidate NPC disease in particular and intracellular sterol transport in general.
|Munkacsi, Andrew B; Hammond, Natalie; Schneider, Remy T et al. (2017) Normalization of Hepatic Homeostasis in the Npc1nmf164 Mouse Model of Niemann-Pick Type C Disease Treated with the Histone Deacetylase Inhibitor Vorinostat. J Biol Chem 292:4395-4410|
|Gulati, Sonia; Balderes, Dina; Kim, Christine et al. (2015) ATP-binding cassette transporters and sterol O-acyltransferases interact at membrane microdomains to modulate sterol uptake and esterification. FASEB J 29:4682-94|
|Ruggles, Kelly V; Garbarino, Jeanne; Liu, Ying et al. (2014) A functional, genome-wide evaluation of liposensitive yeast identifies the ""ARE2 required for viability"" (ARV1) gene product as a major component of eukaryotic fatty acid resistance. J Biol Chem 289:4417-31|
|Ruggles, Kelly V; Turkish, Aaron; Sturley, Stephen L (2013) Making, baking, and breaking: the synthesis, storage, and hydrolysis of neutral lipids. Annu Rev Nutr 33:413-51|
|Sturley, Stephen L; Hussain, M Mahmood (2012) Lipid droplet formation on opposing sides of the endoplasmic reticulum. J Lipid Res 53:1800-10|
|Munkacsi, Andrew B; Chen, Fannie W; Brinkman, Matthew A et al. (2011) An ""exacerbate-reverse"" strategy in yeast identifies histone deacetylase inhibition as a correction for cholesterol and sphingolipid transport defects in human Niemann-Pick type C disease. J Biol Chem 286:23842-51|
|Shechtman, Caryn F; Henneberry, Annette L; Seimon, Tracie A et al. (2011) Loss of subcellular lipid transport due to ARV1 deficiency disrupts organelle homeostasis and activates the unfolded protein response. J Biol Chem 286:11951-9|
|Fakas, Stylianos; Qiu, Yixuan; Dixon, Joseph L et al. (2011) Phosphatidate phosphatase activity plays key role in protection against fatty acid-induced toxicity in yeast. J Biol Chem 286:29074-85|
|Gulati, Sonia; Liu, Ying; Munkacsi, Andrew B et al. (2010) Sterols and sphingolipids: dynamic duo or partners in crime? Prog Lipid Res 49:353-65|
|Madra, Moneek; Sturley, Stephen L (2010) Niemann-Pick type C pathogenesis and treatment: from statins to sugars. Clin Lipidol 5:387-395|
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