My long-term research objective is aimed at verifying the structure and function of cytoskeletal systems in amoeboid cells in terms of the molecular basis of cellular behavior with regard to the dynamic reorganization of cytoskeletal elements during cellular motile processes. The specific goals of the present proposal are to extend our recent findings that myosin molecules form thick filaments in Dictyostelium and its organization is changed in a dynamic manner during the chemotactic response to c-AMP in population of cells. To obtain more direct evidence, we wish to perform a detailed analysis of actomyosin and microtubule systems in individual cells locally stimulated with chemotactic agents using micromanipulators. This line of studies will be also done on individually identified cells using a motion analysis system. We also wish to examine the coordinated changes in these two cytoskeletal systems in individual mitotic cells. Thirdly, we wish to analyze the cytoskeletal organization of mutant cells which include abnormal myosin. To assess the possible role of myosin-I, we also wish to study differential localization of myosin-I and II. Finally, we will attempt to activate permeabilized cells with ATP to test the sliding movement of individual myosin filaments. The reactivating model will be also applied to studying differential functions of myosin-I and II. This proposal also aims at an electron microscopic identification of the myosin filaments using the quick-freeze, deep-etched platinum replication method. The methods will include high-resolution, high-contrast, high-sensitive videomicroscopy coupled with image-processing system, the application of chemotactic agents using ionophoresis apparatus equipped with micromanipulator, analysis of the behavioral response using motion-analysis system, and platinum-replication transmission electron microscopy.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM039548-04
Application #
3296632
Study Section
Cellular Biology and Physiology Subcommittee 1 (CBY)
Project Start
1988-12-01
Project End
1993-11-30
Budget Start
1991-12-01
Budget End
1992-11-30
Support Year
4
Fiscal Year
1992
Total Cost
Indirect Cost
Name
Northwestern University at Chicago
Department
Type
Schools of Dentistry
DUNS #
005436803
City
Chicago
State
IL
Country
United States
Zip Code
60611
Fukui, Yoshio (2002) Mechanistics of amoeboid locomotion: signal to forces. Cell Biol Int 26:933-44
Fukui, Y; de Hostos, E; Yumura, S et al. (1999) Architectural dynamics of F-actin in eupodia suggests their role in invasive locomotion in Dictyostelium. Exp Cell Res 249:33-45
Fukui, Y; Engler, S; Inoue, S et al. (1999) Architectural dynamics and gene replacement of coronin suggest its role in cytokinesis. Cell Motil Cytoskeleton 42:204-17
Fukui, Y; Kitanishi-Yumura, T; Yumura, S (1999) Myosin II-independent F-actin flow contributes to cell locomotion in dictyostelium. J Cell Sci 112 ( Pt 6):877-86
Yumura, S; Fukui, Y (1998) Spatiotemporal dynamics of actin concentration during cytokinesis and locomotion in Dictyostelium. J Cell Sci 111 ( Pt 15):2097-108
Aizawa, H; Fukui, Y; Yahara, I (1997) Live dynamics of Dictyostelium cofilin suggests a role in remodeling actin latticework into bundles. J Cell Sci 110 ( Pt 19):2333-44
Fukui, Y; Inoue, S (1997) Amoeboid movement anchored by eupodia, new actin-rich knobby feet in Dictyostelium. Cell Motil Cytoskeleton 36:339-54
Chu, Q; Fukui, Y (1996) In vivo dynamics of myosin II in Dictyostelium by fluorescent analogue cytochemistry. Cell Motil Cytoskeleton 35:254-68
Morita, Y S; Jung, G; Hammer 3rd, J A et al. (1996) Localization of Dictyostelium myoB and myoD to filopodia and cell-cell contact sites using isoform-specific antibodies. Eur J Cell Biol 71:371-9
Fukui, Y; Inoue, S (1995) Chemotaxis, aggregation behavior, and foot formation in Dictyostelium discoideum amoeba controlled by microbeam uncaging of cyclic-AMP. Biol Bull 189:198-9

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