Dynamics of putative raft-associated proteins at the cell surface.
Lipid rafts are conceptualized as membrane microdomains enriched in cholesterol and glycosphingolipid that serve as platforms for protein segregation and signaling. The properties of these domains in vivo are unclear. Here, we use fluorescence recovery after photobleaching to test if raft association affects a protein's ability to laterally diffuse large distances across the cell surface. The diffusion coefficients (D) of several types of putative raft and nonraft proteins were systematically measured under steady-state conditions and in response to raft perturbations. Raft proteins diffused freely over large distances (> 4 microm), exhibiting Ds that varied 10-fold. This finding indicates that raft proteins do not undergo long-range diffusion as part of discrete, stable raft domains. Perturbations reported to affect lipid rafts in model membrane systems or by biochemical fractionation (cholesterol depletion, decreased temperature, and cholesterol loading) had similar effects on the diffusional mobility of raft and nonraft proteins. Thus, raft association is not the dominant factor in determining long-range protein mobility at the cell surface.
Kenworthy
Anne K
AK
Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bldg. 18T, Rm. 101, 18 Library Dr., Bethesda, MD 20895, USA.
Nichols
Benjamin J
BJ
Remmert
Catha L
CL
Hendrix
Glenn M
GM
Kumar
Mukesh
M
Zimmerberg
Joshua
J
Lippincott-Schwartz
Jennifer
J
D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D002784:Cholesterol / Q000172:deficiency; D006224:Cricetinae; D003505:Cyclodextrins / Q000494:pharmacology; D004058:Diffusion; D036681:Fluorescence Recovery After Photobleaching; D008563:Membrane Lipids / Q000378:metabolism; D021962:Membrane Microdomains / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008954:Models, Biological; D017711:Nonlinear Dynamics; D051381:Rats; D013696:Temperature; D047392:beta-Cyclodextrins
D003505:Cyclodextrins; D008563:Membrane Lipids; D008565:Membrane Proteins; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D002784:Cholesterol