Dynamic regulation of the actin cytoskeleton is important for cell motility, spreading and the formation of membrane surface extensions such as lamellipodia, ruffles and blebs. The ubiquitous calpains contribute to integrin-mediated cytoskeletal remodelling during cell migration and spreading, by cleavage of focal adhesion components and signalling molecules. In the present study, the live-cell morphology of calpain-knockout and wild-type cells was examined by time-lapse fluorescence microscopy, and a role of calpain in mediating the formation of sporadic membrane blebs was established. Membrane blebbing was significantly reduced in calpain-knockout cells, and genetic rescue fully restored the wild-type phenotype in knockout cells. Proteomic comparison of wild-type and knockout cells identified decreased levels of RhoGDI-1 (Rho GDP-dissociation inhibitor) and cofilin 1, and increased levels of tropomyosin in calpain-knockout cells, suggesting a role of calpain in regulating membrane extensions involving these proteins. RhoGDI, cofilin and tropomyosin are known regulators of actin filament dynamics and membrane extensions. The reduced levels of RhoGDI-1 in calpain-knockout cells observed by proteome analysis were confirmed by immunoblotting. Genetic rescue of the calpain-knockout cells enhanced RhoGDI-1-expression 2-fold above that normally present in wild-type cells. These results suggest a regulatory connection between calpain and RhoGDI-1 in promoting formation of membrane blebs.
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Research Article|
April 14 2008
Genetic disruption of calpain correlates with loss of membrane blebbing and differential expression of RhoGDI-1, cofilin and tropomyosin
Anna K. Larsen;
Anna K. Larsen
1
*Department of Food Science, Faculty of Life Sciences, University of Copenhagen, Rolighedsvej 30, DK-1958 Frederiksberg C, Denmark
1To whom correspondence should be addressed (email akl@life.ku.dk).
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René Lametsch;
René Lametsch
*Department of Food Science, Faculty of Life Sciences, University of Copenhagen, Rolighedsvej 30, DK-1958 Frederiksberg C, Denmark
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John S. Elce;
John S. Elce
†Department of Biochemistry, Queen's University, Botterell Hall, Kingston, ON, Canada K7L 3N6
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Jørgen K. Larsen;
Jørgen K. Larsen
‡Finsen Laboratory and Experimental Pathology Unit, Rigshospitalet, Copenhagen Biocenter, Ole Maaløes Vej 5, DK-2200 København N, Denmark
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Bo Thomsen;
Bo Thomsen
§Department of Genetics and Biotechnology, Faculty of Agricultural Science, University of Aarhus, Research Centre Foulum, Blichers Allé 20, DK-8830 Tjele, Denmark
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Martin R. Larsen;
Martin R. Larsen
∥Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense, Denmark
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Moira A. Lawson;
Moira A. Lawson
*Department of Food Science, Faculty of Life Sciences, University of Copenhagen, Rolighedsvej 30, DK-1958 Frederiksberg C, Denmark
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Peter A. Greer;
Peter A. Greer
¶Queen's Cancer Research Institute, Queen's University, Botterell Hall, Room A309, Kingston, ON, Canada K7L 3N6
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Per Ertbjerg
Per Ertbjerg
*Department of Food Science, Faculty of Life Sciences, University of Copenhagen, Rolighedsvej 30, DK-1958 Frederiksberg C, Denmark
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Publisher: Portland Press Ltd
Received:
April 17 2007
Revision Received:
November 29 2007
Accepted:
December 12 2007
Accepted Manuscript online:
December 12 2007
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2008 Biochemical Society
2008
Biochem J (2008) 411 (3): 657–666.
Article history
Received:
April 17 2007
Revision Received:
November 29 2007
Accepted:
December 12 2007
Accepted Manuscript online:
December 12 2007
Citation
Anna K. Larsen, René Lametsch, John S. Elce, Jørgen K. Larsen, Bo Thomsen, Martin R. Larsen, Moira A. Lawson, Peter A. Greer, Per Ertbjerg; Genetic disruption of calpain correlates with loss of membrane blebbing and differential expression of RhoGDI-1, cofilin and tropomyosin. Biochem J 1 May 2008; 411 (3): 657–666. doi: https://doi.org/10.1042/BJ20070522
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