Issue 17, 2014

Imaging lipid order changes in endosome membranes of live cells by using a Nile Red-based membrane probe

Abstract

During maturation of endosomes, in addition to the decrease in their internal pH and the modifications of their proteins, changes are thought to occur at the level of their lipid membranes. In the present work, we used the recently developed environment-sensitive membrane probe NR12S to monitor in living cells the lipid order changes that accompany the maturation of endosomes. Internalization studies in HeLa cells using two-photon fluorescence microscopy in the presence of endocytosis markers and inhibitors show that the probe is endocytosed through different energy-dependent pathways. While marginal changes in the probe colour occur during the initial steps of endocytosis, dramatic changes in colour are observed in the membranes of late endosomes and lysosomes. This remarkable colour change, which takes place 2 h after initial binding of NR12S to cell plasma membranes, suggests a loss of lipid ordered phase during endosome maturation. This change of lipid phase likely results from the decrease in the cholesterol content and the hydrolysis of sphingomyelin occurring in the membranes of late endosomes and lysosomes. In comparison to the common endocytosis marker FM4-64, NR12S is many-fold brighter, it can monitor in situ changes in the lipid organization of endosome membranes and its fluorescence at the plasma membrane can be selectively switched off by sodium dithionite. The present work proposes thus a new powerful tool for endocytosis research that enables monitoring changes in the lipid composition of endosome membranes.

Graphical abstract: Imaging lipid order changes in endosome membranes of live cells by using a Nile Red-based membrane probe

Supplementary files

Article information

Article type
Paper
Submitted
01 Des 2013
Accepted
14 Jan 2014
First published
15 Jan 2014

RSC Adv., 2014,4, 8481-8488

Author version available

Imaging lipid order changes in endosome membranes of live cells by using a Nile Red-based membrane probe

Z. Darwich, A. S. Klymchenko, D. Dujardin and Y. Mély, RSC Adv., 2014, 4, 8481 DOI: 10.1039/C3RA47181K

To request permission to reproduce material from this article, please go to the Copyright Clearance Center request page.

If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given.

If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page.

Read more about how to correctly acknowledge RSC content.

Social activity

Spotlight

Advertisements