J. Lipid Res. Neurobiology of Lipids (ISSN1683-5506)
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH
QUICK SEARCH:   [advanced]


     

A more recent version of this article appeared on May 1, 2003

Papers In Press, published online ahead of print February 1, 2003
J. Lipid Res., doi:10.1194/jlr.D200036-JLR200
This Article
Right arrow Full Text (Accepted Manuscript)
Right arrow All Versions of this Article:
D200036-JLR200v1
44/5/1033    most recent
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Sugii, S.
Right arrow Articles by Chang, T.-Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Sugii, S.
Right arrow Articles by Chang, T.-Y.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

Submitted on September 29, 2002
Revised on December 5, 2002
Accepted on January 28, 2003

Biotinylated theta toxin derivative as a probe to examine intracellular cholesterol-rich domains in normal and Niemann-pick type C1 cells

Shigeki Sugii, Patrick C. Reid, Nobutaka Ohgami, Yukiko Shimada, Robert A. Maue, Haruaki Ninomiya, Yoshiko Ohno-Iwashita, and Ta-Yuan Chang

Department of Biochemistry, Dartmouth Medical School, Hanover, NH 03755

Corresponding Author: Ta.Yuan.Chang{at}Dartmouth.edu

BCtheta is a proteolytically nicked and biotinylated derivative of a cholesterol binding protein perfringolysin O (theta -toxin), and has been used to detect cholesterol-rich domains at the plasma membrane. Here we show that by modifying the cell fixation condition, BCtheta can also be used to detect cholesterol-rich domains intracellularly. When cells were processed for plasma membrane cholesterol staining, the difference in BCtheta signals between the CT43 cell, a mutant Chinese hamster ovary (CHO) cell line lacking the Niemann-Pick C1 (NPC1) protein, and its parental cell 25RA was minimal. However, when cells were fixed with 4% paraformaldehyde, they became permeable to BCtheta . Under this condition, BCtheta mainly stained cholesterol-rich domains inside the cells, with the signal being much stronger in CT43 cells than in 25RA cells. The sensitivity of BCtheta staining was superior to that of filipin staining. The staining of cholesterol-rich domain(s) inside 25RA cells was sensitive to beta -cyclodextrin treatment, while most of the staining inside CT43 cells was relatively resistant to cyclodextrin treatment. Clear differences in intracellular BCtheta staining were also seen between the normal and mutant NPC1 fibroblasts of human or mouse origin. Thus, BCtheta is a powerful tool for visually monitoring cholesterol-rich domains inside normal and NPC cells.


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


This article has been cited by other articles:


Home page
 J. Biol. Chem.Home page
S. Sugii, S. Lin, N. Ohgami, M. Ohashi, C. C. Y. Chang, and T.-Y. Chang
Roles of Endogenously Synthesized Sterols in the Endocytic Pathway
J. Biol. Chem., August 11, 2006; 281(32): 23191 - 23206.
[Abstract] [Full Text] [PDF]

Home page
 J. Cell Biol.Home page
Y. Nagata, T. A. Partridge, R. Matsuda, and P. S. Zammit
Entry of muscle satellite cells into the cell cycle requires sphingolipid signaling
J. Cell Biol., July 17, 2006; 174(2): 245 - 253.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
I. G. Ganley and S. R. Pfeffer
Cholesterol Accumulation Sequesters Rab9 and Disrupts Late Endosome Function in NPC1-deficient Cells
J. Biol. Chem., June 30, 2006; 281(26): 17890 - 17899.
[Abstract] [Full Text] [PDF]

Home page
 Infect. Immun.Home page
R. K. Tweten
Cholesterol-Dependent Cytolysins, a Family of Versatile Pore-Forming Toxins
Infect. Immun., October 1, 2005; 73(10): 6199 - 6209.
[Full Text] [PDF]

Home page
 J BiochemHome page
R. Ishitsuka, S. B. Sato, and T. Kobayashi
Imaging Lipid Rafts
J. Biochem., March 1, 2005; 137(3): 249 - 254.
[Abstract] [Full Text] [PDF]

Home page
 J. Lipid Res.Home page
W. Stockinger, A. B. Castoreno, Y. Wang, J. C. Pagnon, and A. Nohturfft
Real-time analysis of endosomal lipid transport by live cell scintillation proximity assay
J. Lipid Res., November 1, 2004; 45(11): 2151 - 2158.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
S. B. Sato, K. Ishii, A. Makino, K. Iwabuchi, A. Yamaji-Hasegawa, Y. Senoh, I. Nagaoka, H. Sakuraba, and T. Kobayashi
Distribution and Transport of Cholesterol-rich Membrane Domains Monitored by a Membrane-impermeant Fluorescent Polyethylene Glycol-derivatized Cholesterol
J. Biol. Chem., May 28, 2004; 279(22): 23790 - 23796.
[Abstract] [Full Text] [PDF]

Home page
 J. Lipid Res.Home page
P. C. Reid, N. Sakashita, S. Sugii, Y. Ohno-Iwashita, Y. Shimada, W. F. Hickey, and T.-Y. Chang
A novel cholesterol stain reveals early neuronal cholesterol accumulation in the Niemann-Pick type C1 mouse brain
J. Lipid Res., March 1, 2004; 45(3): 582 - 591.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
M. Ohashi, N. Mizushima, Y. Kabeya, and T. Yoshimori
Localization of Mammalian NAD(P)H Steroid Dehydrogenase-like Protein on Lipid Droplets
J. Biol. Chem., September 19, 2003; 278(38): 36819 - 36829.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH
 All ASBMB Journals   Journal of Biological Chemistry 
 Molecular and Cellular Proteomics   ASBMB Today 
Copyright © 2003 by the American Society for Biochemistry and Molecular Biology.