(provide by applicant): The overall goal of this research is to understand the role of protein fatty acylation and membrane cholesterol in directing subcellular localization and function of signal transduction proteins. Members of the Src family of tyrosine protein kinases will be used as model systems. Specifically, the experiments outlined in this application are designed to address the following: ? ? 1. To identify signaling molecules and pathways activated by plasma membrane cholesterol depletion. We recently showed that depletion of membrane cholesterol with cyclodextrin activates MAP kinase and causes increased tyrosine phosphorylation on a number of proteins. The identity of these proteins will be determined, as well as the relationship(s) between MAPK activation and protein tyrosine phosphorylation. The intracellular signaling pathways that are activated will be identified using pharmacological and dominant negative protein inhibitors. ? ? 2. To determine the kinetics and regulation of Src family kinase (SFK) depalmitoylation. Palmitoylation is a dynamic reaction that controls protein localization and signaling. We will determine whether APT1, a newly described thioesterase, depalmitoylates SFKs and whether SFK depalmitoylation is regulated by raft localization and/or signal transduction. ? ? 3. To identify the intracellular trafficking mechanism(s) utilized by Src family kinases. The subcellular trafficking pathway(s) involved in directing SFKs to specific membrane locations will be determined using combinations of confocal microscopy and time-lapse imaging of GFP-tagged SFKs. We will test whether SFKs interact with components of the endocytic machinery en route to the plasma membrane.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Study Section
Special Emphasis Panel (ZRG1-CDF-4 (02))
Program Officer
Chin, Jean
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Sloan-Kettering Institute for Cancer Research
New York
United States
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Matevossian, Armine; Resh, Marilyn D (2015) Hedgehog Acyltransferase as a target in estrogen receptor positive, HER2 amplified, and tamoxifen resistant breast cancer cells. Mol Cancer 14:72
Matevossian, Armine; Resh, Marilyn D (2015) Membrane topology of hedgehog acyltransferase. J Biol Chem 290:2235-43
Petrova, E; Matevossian, A; Resh, M D (2015) Hedgehog acyltransferase as a target in pancreatic ductal adenocarcinoma. Oncogene 34:263-8
Rios-Esteves, Jessica; Haugen, Brittany; Resh, Marilyn D (2014) Identification of key residues and regions important for porcupine-mediated Wnt acylation. J Biol Chem 289:17009-19
Callier, Patrick; Calvel, Pierre; Matevossian, Armine et al. (2014) Loss of function mutation in the palmitoyl-transferase HHAT leads to syndromic 46,XY disorder of sex development by impeding Hedgehog protein palmitoylation and signaling. PLoS Genet 10:e1004340
Resh, Marilyn D (2013) Covalent lipid modifications of proteins. Curr Biol 23:R431-5
Rios-Esteves, Jessica; Resh, Marilyn D (2013) Stearoyl CoA desaturase is required to produce active, lipid-modified Wnt proteins. Cell Rep 4:1072-81
Petrova, Elissaveta; Rios-Esteves, Jessica; Ouerfelli, Ouathek et al. (2013) Inhibitors of Hedgehog acyltransferase block Sonic Hedgehog signaling. Nat Chem Biol 9:247-9
Buglino, John A; Resh, Marilyn D (2012) Palmitoylation of Hedgehog proteins. Vitam Horm 88:229-52
Hardy, Rayshonda Y; Resh, Marilyn D (2012) Identification of N-terminal residues of Sonic Hedgehog important for palmitoylation by Hedgehog acyltransferase. J Biol Chem 287:42881-9

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