Studies of the decline of deoxyribonucleic acid polymerase activity during embryonic muscle cell fusion in vitro

O'Neill, Michael C.; Strohman, Richard C.

doi:10.1021/bi00816a012

Cited by 30 publications

(5 citation statements)

References 25 publications

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“…Withdrawal occurs prior to fusion, and is likely to be an endogenously programmed event that subsequently leads to the restructuring of the cell surface that permits fusion between competent myoblasts. These findings regarding post-mitotic myoblasts render unlikely the claim of O'Neill and Strohman (43) and Stockdale and O'Neill (44) that fusion is responsible for the loss of DNA polymerase activity during myogenesis.…”

Section: Differential Response Of Replicating and Post-mitotic Cells Tomentioning

confidence: 82%

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Effects of cytochaslasin B and colcemide on myogenic cultures.

Holtzer¹,

Croop²,

Dienstman³

et al. 1975

Proc. Natl. Acad. Sci. U.S.A.

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Muscle cultures treated with cytochalasin B yield mono-and oligonucleated cells of two kinds: (i) arborized, replicating precursor myogenic cells and fibroblasts; and (ii) round, post-mitotic, terminally differentiating myoblasts and myotubes. The arb6rized cells do not bind fluorescein-labeled antibody against myosin, do not contract rhythmically, and do not display hexagonally stacked thick and thin filaments. The round, mononucleated myoblasts and round, oligonucleated myotubes bind the fluorescein-labeled antibody against myosin, contract rhythmically, and display clusters of hexagonallystacked thick and thin filaments. When cytochalasin B is removed and replaced by colcemide, the arborized cells, but not the post-mitotic muscle cells, acquire a radial symmetry and are induced to assemble massive, meandering cables that may occupy over 25% of the cell volume. These tortuous cables are positively birefringent and consist exclusively of enormous numbers of 100-A, intermediate-sized filaments.After 3 days in vitro primary cultures of embryonic chick muscle contain immature, post-mitotic, multinucleated myotubes, post-mitotic mononucleated myoblasts, replicating myogenic cells, replicating fibrogenic cells, and replicating mesenchyme cells (1,2,40). The addition of cytochalasin B to these cultures promptly blocks fusion of myoblasts into myotubes. Cytochalasin B also rapidly blocks locomotion of all replicating cells, induces them to "retract," and gradually transforms them into fully arborized cells (3-6). Cytochalasin B does not arborize immature myotubes, but it does cause them to retract from the clot, and subsequently the myotubes degenerate. Because cytocholasin B blocks, not the initiation, but the completion, of cytokinesis (6, 7), these cultures show an increase in oligonucleated cells with time. Cytocholasin B does not significantly impair either the initiation or continued synthesis of myosin, actin, or tropomyosin, for by day 6 the post-mitotic cells generated in these cultures contract spontaneously (3, 6).The addition of colcemide to 3-day-old muscle cultures results in the metaphase-arrest of many mononucleated myogenic and fibrogenic cells. Colcemide fragments the elongated, immature myotubes into multinucleated myosacs and dismantles their microtubules. The myosacs are rich in "lakes" of i00-A, intermediate-sized filaments, and it has been proposed that there is an inverse correlation between the microtubules in myotubes and the 100-A filaments in myosacs (6,8). Colcemide has no detectable effect on the synthesis of contractile proteins, their organization into hexagonally-stacked thick and thin filaments, or their interaction required for spontaneous contractions (9). We wish to report that muscle cultures exposed to cytochalasin B gen-513 erate large numbers of post-mitotic, mononucleated myoblasts rich in scattered stacks of thick and thin filaments. Although Z. H, and I bands are atypical, these scattered stacks of thick and thin filaments permit the cell to contract spontaneously. W...

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Section: Differential Response Of Replicating and Post-mitotic Cells Tomentioning

confidence: 82%

“…The first relates to the claims that fusion causes cells to withdraw from the cell cycle (14)(15)(16)(17)(18). The occurrence of round, post-mitotic, mononucleated myoblasts demonstrates that withdrawal from the cell cycle is not the result of fusion.…”

Section: Differential Response Of Replicating and Post-mitotic Cells Tomentioning

confidence: 99%

Effects of cytochaslasin B and colcemide on myogenic cultures.

Holtzer¹,

Croop²,

Dienstman³

et al. 1975

Proc. Natl. Acad. Sci. U.S.A.

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“…We do not as yet know whether or not more myotubes differentiate automatically due to a larger number of cells at 72 hr, or if NTF affects cell fusion directly. The literature indicates that formation of myotubes in vitro may be a function of both cell density and environment [Bischoff and Holtzer, 1969;Hollenberg et al, 1981;O'Neil and Stockdale, 1972b;O'Neil and Strohman, 1970;Zalin, 19791. In fact, muscle cells seem to be able to condition their environment favoring cell fusion into myotubes [Buckley and Konigsberg, 1974a,b;Doering and Fischman, 1974;Konigsberg, 1971;White et al, 19751. Our finding that fusion of myoblasts does not commence to any large degree prior to 48 hr is consistent with previously published data [Bischoff and Holtzer, 1969;Buckley and Konigsberg, 1974b;O'Neil and Stockdale, 1972al.…”

Section: Kinetics Of Muscle Cell Proliferation and Myogenesismentioning

confidence: 99%

Dose‐dependent initiation of myogenesis by neurotrophic factor

Popiela

Ellis

Festoff

1982

J of Neuroscience Research

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The kinetics of primary chick muscle cells in response to partially purified neurotrophic factor (NTF) were investigated. Chick muscle cells are routinely plated in the absence of NTF but in the presence of horse serum to allow cell attachment to the substratum; a maximum number of cells attaches 5-8 hr after plating, but cells do not proliferate in the absence of NTF, or do so extremely slowly. Subsequent to a routine attachment period of 6 hr, the medium is aspirated and replaced with medium containing 2-20 micrograms/ml of NTF as well as horse serum. Dose responsiveness at elevated NTF concentrations is observed only if F12 medium, supplemented with serum, insulin, transferrin, and selenium is used. In the presence of commercial F10, although supplemented with serum, dose responsiveness is not apparent at more than 2 micrograms/ml of NTF. Upon exposure to NTF, in the presence of supplemented F12 and serum, an 8-hr lag ensues; then, thymidine is incorporated at NTF dose-dependent rates by growing cell populations. The rates of incorporation into cell populations depend on the initial number of cells plated and the amount of NTF supplied. Cell counting indicates that dose-dependent proliferation of muscle cells has occurred during the first 48 hr of exposure to NTF. The specific activity of NTF is repeatedly shown to be at least 10-fold greater than unfractionated nerve extract. Shortly after 48 hr of exposure to NTF, myoblasts characteristically begin to fuse with one another, and myogenesis is visible. These studies show that the initiation of in vitro myogenesis in primary chick muscle cells depends on the amount of NTF supplied. The work also indicates a requirement for optimum medium conditions in order to detect dose-responses to NTF.

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“…Embryonic multinucleated skeletal muscle cells are formed by the fusion of many mononucleated cells which, before cell fusion, divide every 12 hr (1,16) . As the process of differentiation begins, those cells which fuse never again enter the DNA synthetic phase of the cell cycle, and this change is accompanied by a loss of 90% of the DNA polymerase activity coincidentally with cell fusion (24,14,15,22) . However, after UV irradiation, every nucleus (99%) within differentiated muscle cells incorporates tritiated thymidine after UV light treatment (23) .…”

Section: Introductionmentioning

confidence: 99%