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Studies on the Dynamic Localization of GFP-Myosin During Cytokinesis in Live Cells

Published online by Cambridge University Press:  02 July 2020

James Sabry ,
Sheri Moores ,
Shannon Ryan ,
Ji-Hong Zang  and
James A. Spudich
James Sabry
Affiliation:
Department of Biochemistry, Stanford University Medical School, Stanford, CA, 94305
Sheri Moores
Affiliation:
Department of Biochemistry, Stanford University Medical School, Stanford, CA, 94305
Shannon Ryan
Affiliation:
Department of Biochemistry, Stanford University Medical School, Stanford, CA, 94305
Ji-Hong Zang
Affiliation:
Department of Biochemistry, Stanford University Medical School, Stanford, CA, 94305
James A. Spudich
Affiliation:
Department of Biochemistry, Stanford University Medical School, Stanford, CA, 94305
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Extract

Cell division is thought to be powered by the constriction of an actomyosin containing contractile ring found transiently in the cleavage furrow. Conventional myosin II plays a fundamental role in this process of cytokinesis where, in the form of a multimeric complex known as the bipolar thick filament, it is thought to be the molecular motor that generates the force necessary to cause ring constriction.

In order to study the dynamics of this protein in the dividing cell, we have made a fusion protein of the green fluorescent protein (GFP) and the amino terminus of the Dictyostelium myosin heavy chain (GFP-myosin), and imaged the location of this protein in dividing Dictyostelium cells were it is the only myosin II present in the cell. The addition of GFP does not compromise the functioning of the myosin motor as evidenced by the fact that purified GFP-myosin has solution ATPase and in vitro motility kinetics similar to that of non-labelled myosin. In addition, GFP-myosin fully complements the myosin null mutation for both development and cytokinesis in suspension suggesting that GFP-myosin acts as a regulated motor when expressed in cells.

Type
Cell Biology Applications of Green Fluorescent Protein and Other Vital Labeling Probes
Information
Microscopy and Microanalysis , Volume 3 , Issue S2: Proceedings: Microscopy & Microanalysis '97, Microscopy Society of America 55th Annual Meeting, Microbeam Analysis Society 31st Annual Meeting, Histochemical Society 48th Annual Meeting, Cleveland, Ohio, August 10-14, 1997 , August 1997 , pp. 129 - 130
Copyright
Copyright © Microscopy Society of America 1997

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References

1. Rappaport, R., Cytokinesis in Animal Cells, Cambridge U Press, Cambridge, UK, 1996.CrossRefGoogle Scholar

2. Moores, S.L., Sabry, J.H., and Spudich, J.A., Myosin Dynamics in live Dictyostelium cells, Proc Natl Acad Sci USA, 93: 443–446,1996.CrossRefGoogle ScholarPubMed