Date on Master's Thesis/Doctoral Dissertation

8-2016

Document Type

Master's Thesis

Degree Name

M.S.

Department

Pharmacology and Toxicology

Degree Program

Pharmacology and Toxicology, MS

Committee Chair

Ceresa, Brian

Committee Co-Chair (if applicable)

Hood, Joshua

Committee Member

Hood, Joshua

Committee Member

Wattenberg, Brian

Committee Member

Merchant, Michael

Committee Member

Clark, Geoffrey

Author's Keywords

EGFR; endocytosis; signaling; trafficking; subcellular fractionation; early endosome

Abstract

The epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase (RTK) that is an integral component of proliferative signaling. When activated by a ligand at the plasma membrane, EGFR undergoes clathrin-mediated endocytosis. This spatial regulation of the receptor is an important regulator of receptor expression as it mediates its degradation. Endocytosis also has implications on EGFR downstream signaling, though the details are not fully understood. The goal of this thesis is to develop a method to isolate early endosomes in order to study downstream effectors associated with activated EGFR in this compartment. HeLa cells were used to test various subcellular fractionation methods, optimizing each step to develop a protocol that enriches early endosomes. The isolated compartments were then analyzed by mass spectrometry to characterize the protein composition of early endosomes, with the goal of further understanding how the spatial regulation of EGFR affects its downstream signaling.

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