Cell motility and chemotaxis in Dictyostelium amebae lacking myosin heavy chain

Wessels, Deborah; Soll, David R.; Knecht, David A.; Loomis, William F.; Lozanne, Arturo De; Spudich, James A.

doi:10.1016/0012-1606(88)90279-5

Cited by 312 publications

(253 citation statements)

References 18 publications

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“…Wessels et al (32) suggested that myosin II is required for fine tune locomotion during chemotaxis, and therefore cells lacking myosin II fail to exhibit efficient chemotaxis. On the basis of these results, we propose the following model for Dictyostelium chemotaxis.…”

Section: Discussionmentioning

confidence: 99%

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Dictyostelium Myosin II Is Regulated during Chemotaxis by a Novel Protein Kinase C

Abu-Elneel

Karchi

Ravid

1996

Journal of Biological Chemistry

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When cells of Dictyostelium are starved, they acquire the ability to bind cAMP to specific cell surface receptors and to respond to this signal by chemotaxis. The process of chemotaxis involves phosphorylation and reorganization of myosin II (1-5). In response to cAMP stimulation, myosin II, which exists as thick filaments, translocates to the cortex (5). This translocation is correlated with a transient increase in the rate of myosin II heavy chain (MHC) 1 as well as light chain phosphorylation (3, 4, 6). In addition, in vitro studies strongly suggest that MHC phosphorylation plays an important role in the regulation of myosin II filament formation (7-10). The importance of MHC phosphorylation in the regulation of myosin II in vivo was demonstrated by Egelhoff et al. (11). They found that elimination of the MHC phosphorylation sites allows in vivo contractile activity, but this myosin II shows substantial overassembly. Mimicking the negative charge state of phosphorylated myosin II eliminates filament formation in vitro and renders the myosin II unable to drive any tested contractile event in vivo (11).We previously reported the isolation of a MHC-specific PKC (MHC-PKC) from Dictyostelium that phosphorylates Dictyostelium MHC specifically and is homologous to ␣, ␤, and ␥ subtypes of mammalian PKC (9, 12). This kinase, which is membrane-associated and is expressed during Dictyostelium development, was implicated in the increase in MHC phosphorylation during chemotaxis in this species (9). In vitro phosphorylation of MHC by MHC-PKC results in inhibition of myosin II thick filament formation (9), by inducing the formation of a bent monomer of myosin II whose assembly domain is tied up in an intramolecular interaction that precludes intermolecular interaction, which is necessary for thick filament formation (10). The findings that MHC-PKC is a member of the PKC family and that it regulates myosin II assembly suggest a link between the extracellular chemotactic signal and subsequent intracellular events.A considerable amount of information is now available regarding the regulation of myosin II by MHC phosphorylation, and a picture is beginning to emerge in which the molecular changes in myosin II that are involved in MHC phosphorylation are related to cAMP-induced directed cell movement. Nevertheless, the molecular mechanism by which a cAMP signal is transmitted to myosin II, resulting in myosin II localization and chemotaxis, remains unclear. In an attempt to throw some light on this issue, we studied the role of MHC-PKC in vivo by eliminating and by overexpressing the MHC-PKC protein in Dictyostelium cells. Analysis of these cell lines allowed us to address directly the role of MHC-PKC in vivo and consequently the role of MHC phosphorylation in the regulation of myosin II. The results presented here indicate that MHC-PKC is a key modulator in the regulation of myosin II localization in response to the chemoattractant, cAMP.

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Section: Discussionmentioning

confidence: 99%

“…C, membrane-associated MHC-PKC specific activity of wild type and MHC-PKC ϩ cells at the vegetative stage and after 4 h of development. MHC-PKC activity was assayed by incubating purified LMM58 with the MHC-PKC samples in 10 mM Tris (pH 7.6), 6 mM MgCl 2 , 0.2 mM [␥- 32 Ax2 cells (MHC-PKC ϩ cells). Fig.…”

Section: Fig 2 Expression Of Mhc-pkcmentioning

confidence: 99%

Dictyostelium Myosin II Is Regulated during Chemotaxis by a Novel Protein Kinase C

Abu-Elneel

Karchi

Ravid

1996

Journal of Biological Chemistry

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“…Recent molecular genetic experiments on Dictyostelium discoideum show that myosin II is necessary for cytokinesis and multicellular development. Here we use immunofluorescence microscopy with monoclonal and polyclonal antimyosin antibodies to visualize myosin II in cells of the multicellular D. discoideum slug.M YOSIN II is found in all myocytes and in most eukaryote cells (Kom and Hammer, 1988;Warrick and Spudich, 1987) where it is implicated in cytokinesis (Fujiwara and Pollard, 1976;De Lozanne and Spudich, 1987;Knecht and Loomis, 1987; KitanishiYumura and Fukui, 1989), the control of cell shape (Wessels et al ., 1988), the motility of cells (Yumura et al, 1984;Rubino et al, 1984;Spudich and Spudich, 1982 ;Honer, 1988), the maintenance of cell polarity (Fukui et al, 1990), and the capping of membrane receptor proteins (Pasternak et al ., 1989). In multicellular organisms, myosin II may also play an important role in cell-cell interactions .…”

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“…Wessels et al ., 1988 ;Peters et al ., 1988;Manstein et al, 1989) . Atthis point, development is blocked : aggregates of myosin-deficient mutants fail to become mobile-they do not form slugs with normal three-dimensional characteristics (Wessels et al, 1988) . This developmental block may be a result of diminished control over cell shape (Knecht and Loomis,1987 ;Solomon, 1987) .…”

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confidence: 99%

“…These results provide insight into cell motility within a three-dimensional tissue and they are discussed in relation to the possible roles of myosin II in multicellular development . Wessels et al ., 1988 ;Peters et al ., 1988;Manstein et al, 1989) . Atthis point, development is blocked : aggregates of myosin-deficient mutants fail to become mobile-they do not form slugs with normal three-dimensional characteristics (Wessels et al, 1988) .…”

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The distribution of myosin II in Dictyostelium discoideum slug cells.

Eliott

Vardy

Williams

1991

The Journal of Cell Biology

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Abstract. While the role of myosin II in muscle contraction has been well characterized, less is known about the role of myosin II in non-muscle cells. Recent molecular genetic experiments on Dictyostelium discoideum show that myosin II is necessary for cytokinesis and multicellular development. Here we use immunofluorescence microscopy with monoclonal and polyclonal antimyosin antibodies to visualize myosin II in cells of the multicellular D. discoideum slug.M YOSIN II is found in all myocytes and in most eukaryote cells (Kom and Hammer, 1988;Warrick and Spudich, 1987) where it is implicated in cytokinesis (Fujiwara and Pollard, 1976;De Lozanne and Spudich, 1987;Knecht and Loomis, 1987; KitanishiYumura and Fukui, 1989), the control of cell shape (Wessels et al ., 1988), the motility of cells (Yumura et al., 1984;Rubino et al., 1984;Spudich and Spudich, 1982 ;Honer, 1988), the maintenance of cell polarity (Fukui et al., 1990), and the capping of membrane receptor proteins (Pasternak et al ., 1989). In multicellular organisms, myosin II may also play an important role in cell-cell interactions . Studies suggest that actomyosin bands are responsible for folding sheets of epithelia during gastrulation (Odell et al., 1981;Lee et al., 1983) and that myosin II is involved in both mouse morula compaction (Sobel, 1983 ; and resistance to stress in the embryonic chick area opaca (MonnetTschudi and Kucera, 1988) . Other studies involving gene disruption indicate that myosin II is necessary for normal multicellular development in Diciyostelium discoideum (see below) .D. discoideum is a simple, mobile eukaryote that can exist as an amoeba or as a multicellular aggregate (Bonner, 1967;Raper, 1984). D. discoideum thus lends itself to the study of both single cells and three-dimensional tissues. The cytoskeletal proteins ofindividual amoebae have been widely studied (Spudich and Spudich, 1982;Rubino et al., 1984;Fukui et al., 1987;Condeelis et al., 1987;Knecht and Loomis, 1987;De Lozanne and Spudich, 1987;Gerisch et al., 1989), however, those in multicellular aggregates have not .Mutant D. discoideum cells that lack myosin II survive and undergo (albeit slow) amoeboid movement; such cells differentiate into the two types which normally constitute a slug (prestalk and prespore cells) and theyaggregate to form loose mounds ofcells (Knecht and Loomis, 1987 A subpopulation of peripheral and anterior cells label brightly with antimyosin II antibodies, and many of these cells display a polarized intracellular distribution of myosin II. Other cells in the slug label less brightly and their cytoplasm displays a more homogeneous distribution of myosin II. These results provide insight into cell motility within a three-dimensional tissue and they are discussed in relation to the possible roles of myosin II in multicellular development . Wessels et al ., 1988 ;Peters et al ., 1988;Manstein et al., 1989) . Atthis point, development is blocked : aggregates of myosin-deficient mutants fail to become mobile-they do not form slugs with norm...

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