Magnesium stimulation of calcium binding to tubulin and calcium induced depolymerization of microtubules

The activity of a Ca2+-adenosinetriphosphatase (ATP phosphohydrolase, EC 3.6.1.3) increases during the mitotic phase of synchronized mouse mastocytoma P-815X2 cells. The enzyme is synthesized mainly during a distinct period of the interphase and is activated at mitosis. It is thought to regulate the formation of the mitotic apparatus by controlling the concentration of Ca2+ ions at the site of formation of the mitotic spindle.Ca2+ ions have been implicated as regulating factors in mitosis (1). They can interfere with microtubule assembly (2-4) and a system capable of controlling the concentration of these ions at the place of spindle formation has been tentatively identified (1,5).In sea urchins a Ca2+-adenosinetriphosphatase (ATPase; ATP phosphohydrolase, EC 3.6.1.3) exists which shows in early development cyclic fluctuations during the cell cycle; one peak of activity occurs in the first half of the cell cycle and another one at the time of mitosis (6, 7). Whereas the meaning of the first peak is as yet unclear, some evidence has accumulated that the activity increase of the Ca2+-ATPase at mitosis is involved in the regulation of mitosis. If the length of the cell cycle is altered experimentally, the increase of the enzymatic activity still remains linked with mitosis (7,8). In parthenogenetically activated eggs the activity of the enzyme rises whenever a spindle-like figure is formed (5). If only parts of the cell cycle are turned on, which in sea urchins can be performed by NH3 treatment of unfertilized eggs (9, 10), the Ca2+-ATPase cycle is also turned on, reaching its maximum of activity when the chromosomes show a mitosis-like configuration (11).These and the following results suggest that the enzyme is indeed involved in the regulation of mitosis. Therefore, we propose to name this enzyme the "mitotic Ca2+-ATPase."The experiments so far described were carried out on sea urchin eggs, in which cell divisions follow each other without cell growth during interphases. There, fluctuations of an enzyme activity are likely to represent the activation of pre-existing proteins rather than production of additional enzyme molecules. The same Ca2+-ATPase has also been found in growing cells (mouse fibroblasts) and a cyclic change of activity with a peak at mitosis has been observed (12). The purified enzyme promotes the polymerization of brain tubulin onto isolated mitotic apparatus from L cells (12). The interaction of tubulin from nonneuronal cells with isolated spindles has been reported to be enhanced in the presence of the enzyme (13). In the present work we measure the synthesis and the activity of the mitotic Ca2+-ATPase through the entire cell cycle in synchronized mouse mastocytoma cells. The synthesis of new enzyme proteins takes place during interphase. The peak of the enzymatic activity occurs during mitosis, as in sea urchin, eggs, and must reflect the activation of preexisting enzyme molecules. 1610MATERIAL AND METHODS Cells. Mouse mastocytoma P-815X2 cells (14) were grown in medium I supple...

“…4B shows that the Ca2+-ATPase from mouse cells and the one from isolated mitotic apparatus from sea urchins have identical electrophoretic mobilities. (2,3,25).…”

Section: Resultsmentioning

confidence: 99%

“…They can interfere with microtubule assembly (2)(3)(4) and a system capable of controlling the concentration of these ions at the place of spindle formation has been tentatively identified (1,5).…”

mentioning

confidence: 99%

Synthesis and activation of mitotic Ca2+-adenosinetriphosphatase during the cell cycle of mouse mastocytoma cells.

Petzelt¹,

Auel²

1977

“…The concentration of Ca2' required to inhibit microtubule assembly in vitro apparently depends on the purification protocol used. Assembly in crude brain extracts is inhibited by near micromolar Ca2+ (1,5,6), whereas the assembly of purified protein is sensitive to Ca2+ concentrations in the millimolar range only (7). Interestingly, the sensitivity to micromolar Ca2+ of purified microtubule protein can be restored by a factor present in crude brain extracts (8).…”

mentioning

confidence: 88%

Calcium lability of cytoplasmic microtubules and its modulation by microtubule-associated proteins

Schliwa

Euteneuer

Bulinski

et al. 1981

220

100

“…Several of these results indicated that CDR was concentrated in the chromosome-to-pole region of the metaphase-anaphase mitotic apparatus in all cell lines examined (8). This observation, coupled with the-fact that Ca2+ depolymerizes microtubules in vitro (9)(10)(11)(12)(13) and has been implicated in microtubule regulation in vivo (14)(15)(16), suggested that CDR might be involved in Ca2+-dependent depolymerization of microtubules during anaphase chromosome movement. Additionally, the recent observation that the Ca2+ sensitivity of microtubule assembly…”

mentioning

confidence: 95%

“…Although the molecular mechanism of Ca2+-binding protein-dependent inhibition of microtubule assembly remains to be determined, the data of (13) and others (43), and the millimolar Ca2+ sensitivity reported by OhLmsted and ]Borisy (12). There is insufficient data available to determine if the factor described by Nishida and Sakai (17) …”

mentioning

confidence: 99%

Control of microtubule assembly-disassembly by calcium-dependent regulator protein.

Marcum¹,

Dedman²,

Brinkley³

et al. 1978

434

170

The Ca2+-dependent regulator (CDR) protein of cyclic nucleotide phosphodiesterase is a low molecular weight, acidic, Ca2+-binding protein which has been implicated in a number of Ca2+ dependent enzymatic functions. Indirect immunofluorescence has revealed that CDR is specifically associated with the chromosome-to-pole region of the mitotic apparatus during metaphase-anaphase in a pattern distinctly different from that of similar cultured cells stained with anti-