The Spatiotemporal Pattern of Src Activation at Lipid Rafts Revealed by Diffusion-Corrected FRET Imaging

Title
The Spatiotemporal Pattern of Src Activation at Lipid Rafts Revealed by Diffusion-Corrected FRET Imaging
Authors
Shaoying LuMingxing Ouyang성지혜Jin ZhangShu ChienYingxiao Wang
Issue Date
2008-07
Publisher
PLoS computational biology
Citation
VOL 4, NO 7, e1000127-1-e1000127-12
Abstract
Genetically encoded biosensors based on fluorescence resonance energy transfer (FRET) have been widely applied to visualize the molecular activity in live cells with high spatiotemporal resolution. However, the rapid diffusion of biosensor proteins hinders a precise reconstruction of the actual molecular activation map. Based on fluorescence recovery after photobleaching (FRAP) experiments, we have developed a finite element (FE) method to analyze, simulate, and subtract the diffusion effect of mobile biosensors. This method has been applied to analyze the mobility of Src FRET biosensors engineered to reside at different subcompartments in live cells. The results indicate that the Src biosensor located in the cytoplasm moves 4–8 folds faster (0.93±0.06 µm2/sec) than those anchored on different compartments in plasma membrane (at lipid raft: 0.11±0.01 µm2/sec and outside: 0.18±0.02 µm2/sec). The mobility of biosensor at lipid rafts is slower than that outside of lipid rafts and is dominated by two-dimensional diffusion. When this diffusion effect was subtracted from the FRET ratio images, high Src activity at lipid rafts was observed at clustered regions proximal to the cell periphery, which remained relatively stationary upon epidermal growth factor (EGF) stimulation. This result suggests that EGF induced a Src activation at lipid rafts with well-coordinated spatiotemporal patterns. Our FE-based method also provides an integrated platform of image analysis for studying molecular mobility and reconstructing the spatiotemporal activation maps of signaling molecules in live cells.
URI
http://pubs.kist.re.kr/handle/201004/49259
ISSN
1553734X
Appears in Collections:
KIST Publication > Article
Files in This Item:
There are no files associated with this item.
Export
RIS (EndNote)
XLS (Excel)
XML


qrcode

Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.

BROWSE