Endocytosis and intracellular transport of ricin were studied in stable transfected HeLa cells where overexpression of wild-type (WT) or mutant dynamin is regulated by tetracycline. Overexpression of the temperature-sensitive mutant dynG273D at the nonpermissive temperature or the dynK44A mutant inhibits clathrin-dependent endocytosis (Damke, H., T. Baba, A.M. van der Blieck, and S.L. Schmid. 1995. J. Cell Biol. 131: 69–80; Damke, H., T. Baba, D.E. Warnock, and S.L. Schmid. 1994. J. Cell Biol. 127:915–934). Under these conditions, ricin was endocytosed at a normal level. Surprisingly, overexpression of both mutants made the cells less sensitive to ricin. Butyric acid and trichostatin A treatment enhanced dynamin overexpression and increased the difference in toxin sensitivity between cells with normal and mutant dynamin. Intoxication with ricin seems to require toxin transport to the Golgi apparatus (Sandirg, K., and B. van Deurs. 1996. Physiol. Rev. 76:949–966), and this process was monitored by measuring the incorporation of radioactive sulfate into a modified ricin molecule containing a tyrosine sulfation site. The sulfation of ricin was much greater in cells expressing dynWT than in cells expressing dynK44A. Ultrastructural analysis using a ricin-HRP conjugate confirmed that transport to the Golgi apparatus was severely inhibited in cells expressing dynK44A. In contrast, ricin transport to lysosomes as measured by degradation of 125I-ricin was essentially unchanged in cells expressing dynK44A. These data demonstrate that although ricin is internalized by clathrin-independent endocytosis in cells expressing mutant dynamin, there is a strong and apparently selective inhibition of ricin transport to the Golgi apparatus. Also, in cells with mutant dynamin, there is a redistribution of the mannose-6-phosphate receptor.
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9 February 1998
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February 09 1998
Expression of Mutant Dynamin Inhibits Toxicity and Transport of Endocytosed Ricin to the Golgi Apparatus
Alicia Llorente,
Alicia Llorente
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
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Andrzej Rapak,
Andrzej Rapak
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
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Sandra L. Schmid,
Sandra L. Schmid
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
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Bo van Deurs,
Bo van Deurs
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
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Kirsten Sandvig
Kirsten Sandvig
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
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Alicia Llorente
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
Andrzej Rapak
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
Sandra L. Schmid
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
Bo van Deurs
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
Kirsten Sandvig
*Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway; ‡Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037; and §Structural Cell Biology Unit, Department of Anatomy, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen N, Denmark
Address all correspondence to Kirsten Sandvig, Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway. Tel.: 47 22934294. Fax: 47 22508692. E-mail: [email protected]
Received:
June 04 1997
Revision Received:
December 02 1997
Online ISSN: 1540-8140
Print ISSN: 0021-9525
1998
J Cell Biol (1998) 140 (3): 553–563.
Article history
Received:
June 04 1997
Revision Received:
December 02 1997
Citation
Alicia Llorente, Andrzej Rapak, Sandra L. Schmid, Bo van Deurs, Kirsten Sandvig; Expression of Mutant Dynamin Inhibits Toxicity and Transport of Endocytosed Ricin to the Golgi Apparatus . J Cell Biol 9 February 1998; 140 (3): 553–563. doi: https://doi.org/10.1083/jcb.140.3.553
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