Sorting of an internalized plasma membrane lipid between recycling and degradative pathways in normal and Niemann-Pick, type A fibroblasts

doi:10.1083/jcb.111.2.429

This Article

	Full Text (PDF, 5476K)
	Alert me when this article is cited
	Citation Map

Services

	Email this article
	Similar articles in this journal
	Similar articles in PubMed
	Alert me to new content in the JCB
	Download to citation manager

Citing Articles

	Citing Articles via HighWire
	Citing Articles via CrossRef
	Citing Articles via Google Scholar

Google Scholar

	Articles by Koval, M.
	Articles by Pagano, R. E.
	Search for Related Content

PubMed

	PubMed Citation
	Articles by Koval, M.
	Articles by Pagano, R. E.


Social Bookmarking

	What's this?

ARTICLES

Sorting of an internalized plasma membrane lipid between recycling and degradative pathways in normal and Niemann-Pick, type A fibroblasts

M Koval and RE Pagano
Carnegie Institution of Washington, Department of Embryology, Baltimore, Maryland 21210.

We examined the metabolism and intracellular transport of a fluorescent sphingomyelin analogue, N-(N-[6-[(7-nitrobenz-2-oxa-1,3-diazol-4- yl)amino]caproyl])- sphingosylphosphorylcholine (C6-NBD-SM), in both normal and Niemann-Pick, type A (NP-A) human skin fibroblast monolayers. C6-NBD-SM was integrated into the plasma membrane bilayer by transfer of C6-NBD-SM monomers from liposomes to cells at 7 degrees C. The cells were washed, and within 3 min of warming to 37 degrees C, both normal and NP-A fibroblasts had internalized C6-NBD-SM from the plasma membrane, resulting in a punctate pattern of intracellular fluorescence. Rates for C6-NBD-SM internalization and transport from intracellular compartments to the plasma membrane (recycling) were similar for normal and NP-A cells. With increasing time at 37 degrees C, internalized C6-NBD-SM accumulated in the lysosomes of NP-A fibroblasts, while normal fibroblasts showed increasing Golgi apparatus fluorescence with no observable lysosomal labeling. Since NP-A fibroblasts lack lysosomal (acid) sphingomyelinase (A-SMase), this result suggested that hydrolysis of C6-NBD-SM prevented its accumulation in the lysosomes of normal fibroblasts during its transport along the degradative pathway. We used the amount of C6-NBD- SM hydrolysis by A-SMase in normal cells as a measure of C6-NBD-SM transported from the cell surface to the lysosomes. After a lag period, C6-NBD-SM was delivered to the lysosomes at a rate of approximately 8%/h. This rate was approximately 18-19 fold slower than the rate of C6- NBD-SM recycling from intracellular compartments to the plasma membrane. Thus, small amounts of C6-NBD-SM were transported along the degradative pathway, while most endocytosed C6-NBD-SM was sorted for transport along the plasma membrane recycling pathway.
Add to CiteULike CiteULike Add to Complore Complore Add to Connotea Connotea Add to Del.icio.us Del.icio.us Add to Digg Digg Facebook Add to Reddit Reddit Add to Technorati Technorati Twitter What's this?

This article has been cited by other articles:

Deevska, G. M., Rozenova, K. A., Giltiay, N. V., Chambers, M. A., White, J., Boyanovsky, B. B., Wei, J., Daugherty, A., Smart, E. J., Reid, M. B., Merrill, A. H. Jr., Nikolova-Karakashian, M. (2009). Acid Sphingomyelinase Deficiency Prevents Diet-induced Hepatic Triacylglycerol Accumulation and Hyperglycemia in Mice. J. Biol. Chem. 284: 8359-8368 [Abstract] [Full Text]
Koivusalo, M., Jansen, M., Somerharju, P., Ikonen, E. (2007). Endocytic Trafficking of Sphingomyelin Depends on Its Acyl Chain Length. Mol. Biol. Cell 18: 5113-5123 [Abstract] [Full Text]
Sagiv, Y., Hudspeth, K., Mattner, J., Schrantz, N., Stern, R. K., Zhou, D., Savage, P. B., Teyton, L., Bendelac, A. (2006). Cutting Edge: Impaired Glycosphingolipid Trafficking and NKT Cell Development in Mice Lacking Niemann-Pick Type C1 Protein. J. Immunol. 177: 26-30 [Abstract] [Full Text]
Choudhury, A., Sharma, D. K., Marks, D. L., Pagano, R. E. (2004). Elevated Endosomal Cholesterol Levels in Niemann-Pick Cells Inhibit Rab4 and Perturb Membrane Recycling. Mol. Biol. Cell 15: 4500-4511 [Abstract] [Full Text]
Lada, A. T., Davis, M., Kent, C., Chapman, J., Tomoda, H., Omura, S., Rudel, L. L. (2004). Identification of ACAT1- and ACAT2-specific inhibitors using a novel, cell-based fluorescence assay: individual ACAT uniqueness. J. Lipid Res. 45: 378-386 [Abstract] [Full Text]
Puri, V., Jefferson, J. R., Singh, R. D., Wheatley, C. L., Marks, D. L., Pagano, R. E. (2003). Sphingolipid Storage Induces Accumulation of Intracellular Cholesterol by Stimulating SREBP-1 Cleavage. J. Biol. Chem. 278: 20961-20970 [Abstract] [Full Text]
Johnson, A. O., Lampson, M. A., McGraw, T. E. (2001). A Di-Leucine Sequence and a Cluster of Acidic Amino Acids Are Required for Dynamic Retention in the Endosomal Recycling Compartment of Fibroblasts. Mol. Biol. Cell 12: 367-381 [Abstract] [Full Text]
Tepper, A. D., Ruurs, P., Wiedmer, T., Sims, P. J., Borst, J., van Blitterswijk, W. J. (2000). Sphingomyelin Hydrolysis to Ceramide during the Execution Phase of Apoptosis Results from Phospholipid Scrambling and Alters Cell-Surface Morphology. JCB 150: 155-164 [Abstract] [Full Text]
Hao, M., Maxfield, F. R. (2000). Characterization of Rapid Membrane Internalization and Recycling. J. Biol. Chem. 275: 15279-15286 [Abstract] [Full Text]
Forestier, C., Moreno, E., Pizarro-Cerda, J., Gorvel, J.-P. (1999). Lysosomal Accumulation and Recycling of Lipopolysaccharide to the Cell Surface of Murine Macrophages, an In Vitro and In Vivo Study. J. Immunol. 162: 6784-6791 [Abstract] [Full Text]
Chen, C.-S., Bach, G., Pagano, R. E. (1998). Abnormal transport along the lysosomal pathway in Mucolipidosis, type IV disease. Proc. Natl. Acad. Sci. USA 95: 6373-6378 [Abstract] [Full Text]
Yamaji, A., Sekizawa, Y., Emoto, K., Sakuraba, H., Inoue, K., Kobayashi, H., Umeda, M. (1998). Lysenin, a Novel Sphingomyelin-specific Binding Protein. J. Biol. Chem. 273: 5300-5306 [Abstract] [Full Text]
Duan, H., Lin, C.-Y., Mazzone, T. (1997). Degradation of Macrophage ApoE in a Nonlysosomal Compartment. REGULATION BY STEROLS. J. Biol. Chem. 272: 31156-31162 [Abstract] [Full Text]
French, A. R., Tadaki, D. K., Niyogi, S. K., Lauffenburger, D. A. (1995). Intracellular Trafficking of Epidermal Growth Factor Family Ligands Is Directly Influenced by the pH Sensitivity of the Receptor/Ligand Interaction. J. Biol. Chem. 270: 4334-4340 [Abstract] [Full Text]
Thilo, L, Stroud, E, Haylett, T (1995). Maturation of early endosomes and vesicular traffic to lysosomes in relation to membrane recycling. J. Cell Sci. 108: 1791-1803 [Abstract]
Kok, J. W., Hoekstra, K., Eskelinen, S., Hoekstra, D. (1992). Recycling pathways of glucosylceramide in BHK cells: distinct involvement of early and late endosomes. J. Cell Sci. 103: 1139-1152 [Abstract]
Puri, V., Watanabe, R., Singh, R. D., Dominguez, M., Brown, J. C., Wheatley, C. L., Marks, D. L., Pagano, R. E. (2001). Clathrin-dependent and -independent internalization of plasma membrane sphingolipids initiates two Golgi targeting pathways. JCB 154: 535-548 [Abstract] [Full Text]