HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 282, Issue 50, 36682-36691, December 14, 2007

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 282/50/36682    most recent M702959200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Chichili, G. R. Articles by Rodgers, W. Search for Related Content PubMed PubMed Citation Articles by Chichili, G. R. Articles by Rodgers, W. Social Bookmarking                      What's this?

Clustering of Membrane Raft Proteins by the Actin Cytoskeleton^*

Gurunadh R. Chichili and William Rodgers^¶1

From the Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma 73104 and the Departments of Microbiology and Immunology and ^¶Pathology, Oklahoma University Health Sciences Center, Oklahoma City, Oklahoma 73104

Cell membranes are laterally organized into functionally discrete domains that include the cholesterol-dependent membrane "rafts." However, how membrane domains are established and maintained remains unresolved and controversial but often requires the actin cytoskeleton. In this study, we used fluorescence resonance energy transfer to measure the role of the actin cytoskeleton in the co-clustering of membrane raft-associated fluorescent proteins (FPs) and FPs targeted to the nonraft membrane fraction. By fitting the fluorescence resonance energy transfer data to an isothermal binding equation, we observed a specific co-clustering of raft-associated donor and acceptor probes that was sensitive to latrunculin B (Lat B), which disrupts the actin cytoskeleton. Conversely, treating with jasplakinolide to enhance actin polymerization increased co-clustering of the raft-associated FPs over that of the nonraft probes. We also observed by immunoblotting experiments that the actin-dependent co-clustering coincided with regulation of the raft-associated Src family kinase Lck. Specifically, Lat B decreased the phosphorylation of the C-terminal regulatory tyrosine of Lck (Tyr⁵⁰⁵), and combining the Lat B with filipin further decreased the Tyr⁵⁰⁵ phosphorylation. Furthermore, the Lat B-dependent changes in Lck regulation required CD45 because no significant changes occurred in treated T cells lacking CD45 expression. These data define a role for the actin cytoskeleton in promoting co-clustering of raft-associated proteins and show that this property is important toward regulating raft-associated signaling proteins such as Lck.

Received for publication, April 9, 2007 , and in revised form, October 10, 2007.

^* This work was supported by National Institutes of Health Grant R01 GM070001. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Fig. S1.

¹ To whom correspondence should be addressed: MS 45, OK Medical Research Foundation, 825 N.E. 13th St., Oklahoma City, OK 73104. Tel.: 405-271-3550; Fax: 405-271-7417; E-mail: william-rodgers{at}omrf.org.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				D.-A. Persaud-Sawin, S. Lightcap, and G. J. Harry Isolation of rafts from mouse brain tissue by a detergent-free method J. Lipid Res., April 1, 2009; 50(4): 759 - 767. [Abstract] [Full Text] [PDF]

				B. Bhattacharya and P. Roy Bluetongue Virus Outer Capsid Protein VP5 Interacts with Membrane Lipid Rafts via a SNARE Domain J. Virol., November 1, 2008; 82(21): 10600 - 10612. [Abstract] [Full Text] [PDF]

				C. M. Johnson, G. R. Chichili, and W. Rodgers Compartmentalization of Phosphatidylinositol 4,5-Bisphosphate Signaling Evidenced Using Targeted Phosphatases J. Biol. Chem., October 31, 2008; 283(44): 29920 - 29928. [Abstract] [Full Text] [PDF]