Quantitative analysis and modulation of the spatial operation of the EGF receptor autocrine cell signaling system

The spatial localization of EGFR autocrine systems is a critical factor for tissue development and homeostasis. It is difficult to analyze these systems, however, because they are both self-contained and recursive. To understand how cellular parameters influence autocrine activity, we investigated a human epidermal growth factor/epidermal growth factor receptor (EGF/EGFR) loop engineered into mouse B82 fibroblasts. We modulated three parameters: ligand secretion, receptor expression and ligand/receptor binding affinity using various molecular and cell biology techniques. We measured autocrine system localization via autocrine ligand release using two different assays: the first indirectly measured the fraction of autocrine ligand captured by measuring ligand escaping the cells and the second measured receptor activation using a microphysiometer-based assay. We analyzed our data using a quantitative model of autocrine cell receptor/ligand dynamics. We found that the activity of our autocrine system could be described in terms of a simple ratio between the rate of ligand production (VLT) and the rate of receptor production (VR). At a VLTNR ratio of < 0.3, essentially no ligand was found in the extracellular medium, but a significant number cell receptors (30-40%) were occupied. As the VLTNR ratio increased from 0.3 towards unity, receptor occupancy increased, and significant amounts of ligand appeared in the medium.

Department dc:contributor.department

Massachusetts Institute of Technology. Dept. of Chemical Engineering.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2001

Author dc:creator

• DeWitt, Anne Elizabeth, 1974-

Advisor dc:contributor.advisor

• Douglas A. Lauffenburger.

• Chemical Engineering.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/8204

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/8204