A mechanism for the intracellular localization of myosin II filaments in the Dictyostelium amoeba

1993 ◽  
Vol 105 (1) ◽  
pp. 233-242 ◽  
Author(s):  
S. Yumura ◽  
T. Kitanishi-Yumura
Keyword(s):  
Intracellular Ph ◽  
Heavy Chain ◽  
Intracellular Localization ◽  
Myosin Ii ◽  
Cortical Region ◽  
Actin Network ◽  
Vegetative Cells ◽  
Heavy Chains ◽  
Ph Values ◽  
Intracellular Concentrations

When ATP is added to membrane-cytoskeletons prepared from Dictyostelium amoebae by the method described previously (S. Yumura and T. Kitanishi-Yumura, Cell Struct. Funct. 15, 355–364, 1990), myosin II is released from the membrane-cytoskeletons after contraction. Simultaneously, the heavy chains of myosin II are phosphorylated by a putative myosin II heavy-chain kinase, at foci within the actin network, with the resultant disassembly of filaments. In this study, we examined factors that control the release of myosin II from the membrane-cytoskeletons, on the assumption that inhibition of the release of myosin II keeps the myosin II in the cortical region, and is responsible for the localization of myosin II in the cortical region. The release of myosin II is inhibited at pH values below 6.5. This effect is not due to the inhibition of heavy-chain phosphorylation but is due to the suppression of disassembly of the filaments. In the membrane-cytoskeletons of aggregating cells, the release of myosin II is inhibited by Ca2+, and this effect is enhanced by pretreatment with calmodulin. In the membrane-cytoskeletons of vegetative cells, the release of myosin II is inhibited by pretreatment with calmodulin, and this effect is Ca2+-independent. The inhibition of the release of myosin II by Ca2+ and/or calmodulin is due to the inhibition of heavy-chain phosphorylation, and calmodulin is associated with the foci within the actin network. These results represent a possible mechanism for the intracellular localization of myosin II via regulation of the release of myosin from the cortical region by changes in intracellular pH and/or intracellular concentrations of Ca2+.

scholarly journals Release of myosin II from the membrane-cytoskeleton of Dictyostelium discoideum mediated by heavy-chain phosphorylation at the foci within the cortical actin network

1992 ◽  
Vol 117 (6) ◽  
pp. 1231-1239 ◽  
Author(s):  
S Yumura ◽  
T Kitanishi-Yumura
Keyword(s):  
Heavy Chain ◽  
Myosin Ii ◽  
Cortical Region ◽  
Actin Network ◽  
Cortical Actin ◽  
Filamentous Form ◽  
Heavy Chains ◽  
Membrane Cytoskeleton ◽  
Myosin Filaments ◽  
Gamma S

Membrane-cytoskeletons were prepared from Dictyostelium amebas, and networks of actin and myosin II filaments were visualized on the exposed cytoplasmic surfaces of the cell membranes by fluorescence staining (Yumura, S., and T. Kitanishi-Yumura. 1990. Cell Struct. Funct. 15:355-364). Addition of ATP caused contraction of the cytoskeleton with aggregation of part of actin into several foci within the network, but most of myosin II was released via the foci. However, in the presence of 10 mM MgCl2, which stabilized myosin II filaments, myosin II remained at the foci. Ultrastructural examination revealed that, after contraction, only traces of monomeric myosin II remained at the foci. By contrast, myosin II filaments remained in the foci in the presence of 10 mM MgCl2. These observations suggest that myosin II was released not in a filamentous form but in a monomeric form. Using [gamma 32P]ATP, we found that the heavy chains of myosin II released from membrane-cytoskeletons were phosphorylated, and this phosphorylation resulted in disassembly of myosin filaments. Using ITP (a substrate for myosin II ATPase) and/or ATP gamma S (a substrate for myosin II heavy-chain kinase [MHCK]), we demonstrated that phosphorylation of myosin heavy chains occurred at the foci within the actin network, a result that suggests that MHCK was localized at the foci. These results together indicate that, during contraction, the heavy chains of myosin II that have moved toward the foci within the actin network are phosphorylated by a specific MHCK, with the resultant disassembly of filaments which are finally released from membrane-cytoskeletons. This series of reactions could represent the mechanism for the relocation of myosin II from the cortical region to the endoplasm.

scholarly journals Identification of three phosphorylation sites on each heavy chain of Acanthamoeba myosin II.

1981 ◽  
Vol 256 (24) ◽  
pp. 12811-12816 ◽  
Author(s):  
G.P. Côté ◽  
J.H. Collins ◽  
E.D. Korn
Keyword(s):  
Heavy Chain ◽  
Myosin Ii ◽  
Phosphorylation Sites

Construction of Polysiloxane-Based Fluorescent Probe for Visualizing The pH Down-regulation

2021 ◽  
Author(s):  
Xiaoni Wang ◽  
Zhiming Gou ◽  
Minggang Tian ◽  
Yujing Zuo
Keyword(s):  
Fluorescent Probe ◽  
Intracellular Ph ◽  
Cell Apoptosis ◽  
Pathological Process ◽  
Down Regulation ◽  
Ph Values

Intracellular pH values change is a significant physiological and pathological process and plays vital roles in autophagy, self-repairing, and programmed cell apoptosis. A unique fluorescent probe (PN-1) based on polysiloxanes...

Quadruply-labeled serum albumin as a biodegradable nanosensor for simultaneous fluorescence imaging of intracellular pH values, oxygen and temperature

2019 ◽  
Vol 186 (8) ◽  
Author(s):  
Xiao-ai Zhang ◽  
Wei Zhang ◽  
Qi Wang ◽  
Junli Wang ◽  
Guodong Ren ◽  
...  
Keyword(s):  
Serum Albumin ◽  
Fluorescence Imaging ◽  
Intracellular Ph ◽  
Ph Values

scholarly journals Novel Myosin Heavy Chain Kinase Involved in Disassembly of Myosin II Filaments and Efficient Cleavage in MitoticDictyostelium Cells

2002 ◽  
Vol 13 (12) ◽  
pp. 4333-4342 ◽  
Author(s):  
Akira Nagasaki ◽  
Go Itoh ◽  
Shigehiko Yumura ◽  
Taro Q.P. Uyeda
Keyword(s):  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Myosin Ii ◽  
Kinase Domain ◽  
Cleavage Furrow ◽  
Wild Type ◽  
Daughter Cells ◽  
Cleavage Process ◽  
Interphase Cells ◽  
Myosin Heavy Chain Kinase

We have cloned a full-length cDNA encoding a novel myosin II heavy chain kinase (mhckC) from Dictyostelium. Like other members of the myosin heavy chain kinase family, themhckC gene product, MHCK C, has a kinase domain in its N-terminal half and six WD repeats in the C-terminal half. GFP-MHCK C fusion protein localized to the cortex of interphase cells, to the cleavage furrow of mitotic cells, and to the posterior of migrating cells. These distributions of GFP-MHCK C always corresponded with that of myosin II filaments and were not observed in myosin II-null cells, where GFP-MHCK C was diffusely distributed in the cytoplasm. Thus, localization of MHCK C seems to be myosin II-dependent. Cells lacking the mhckC gene exhibited excessive aggregation of myosin II filaments in the cleavage furrows and in the posteriors of the daughter cells once cleavage was complete. The cleavage process of these cells took longer than that of wild-type cells. Taken together, these findings suggest MHCK C drives the disassembly of myosin II filaments for efficient cytokinesis and recycling of myosin II that occurs during cytokinesis.

scholarly journals The effect of heavy chain phosphorylation and solution conditions on the assembly of Acanthamoeba myosin-II.

1989 ◽  
Vol 109 (4) ◽  
pp. 1529-1535 ◽  
Author(s):  
J H Sinard ◽  
T D Pollard
Keyword(s):  
Light Scattering ◽  
Ionic Strength ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Critical Concentration ◽  
Myosin Ii ◽  
Acidic Ph ◽  
Thick Filaments ◽  
Low Ionic Strength

At low ionic strength, Acanthamoeba myosin-II polymerizes into bipolar minifilaments, consisting of eight molecules, that scatter about three times as much light as monomers. With this light scattering assay, we show that the critical concentration for assembly in 50-mM KCl is less than 5 nM. Phosphorylation of the myosin heavy chain over the range of 0.7 to 3.7 P per molecule has no effect on its KCl dependent assembly properties: the structure of the filaments, the extent of assembly, and the critical concentration for assembly are the same. Sucrose at a concentration above a few percent inhibits polymerization. Millimolar concentrations of MgCl2 induce the lateral aggregation of fully formed minifilaments into thick filaments. Compared with dephosphorylated minifilaments, minifilaments of phosphorylated myosin have a lower tendency to aggregate laterally and require higher concentrations of MgCl2 for maximal light scattering. Acidic pH also induces lateral aggregation, whereas basic pH leads to depolymerization of the myosin-II minifilaments. Under polymerizing conditions, millimolar concentrations of ATP only slightly decrease the light scattering of either phosphorylated or dephosphorylated myosin-II. Barring further modulation of assembly by unknown proteins, both phosphorylated and dephosphorylated myosin-II are expected to be in the form of minifilaments under the ionic conditions existing within Acanthamoeba.

Intracellular pH homeostasis during cell-cycle progression and growth state transition in Schizosaccharomyces pombe

2001 ◽  
Vol 114 (16) ◽  
pp. 2929-2941 ◽  
Author(s):  
Jim Karagiannis ◽  
Paul G. Young
Keyword(s):  
Cell Cycle ◽  
Schizosaccharomyces Pombe ◽  
Intracellular Ph ◽  
Cell Cycle Progression ◽  
State Transition ◽  
Homeostatic Regulation ◽  
Ph Homeostasis ◽  
Vegetative Cells ◽  
Growth State ◽  
Internal Ph

Accurate measurement of intracellular pH in unperturbed cells is fraught with difficulty. Nevertheless, using a variety of methods, intracellular pH oscillations have been reported to play a regulatory role in the control of the cell cycle in several eukaryotic systems. Here, we examine pH homeostasis in Schizosaccharomyces pombe using a non-perturbing ratiometric pH sensitive GFP reporter. This method allows for accurate intracellular pH measurements in living, entirely undisturbed, logarithmically growing cells. In addition, the use of a flow cell allows internal pH to be monitored in real time during nutritional, or growth state transition. We can find no evidence for cell-cycle-related changes in intracellular pH. By contrast, all data are consistent with a very tight homeostatic regulation of intracellular pH near 7.3 at all points in the cell cycle. Interestingly, pH set point changes are associated with growth state. Spores, as well as vegetative cells starved of either nitrogen, or a carbon source, show a marked reduction in their internal pH compared with logarithmically growing vegetative cells. However, in both cases, homeostatic regulation is maintained.

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