On the ‘clock’ mechanism determining the time of tissue-specific enzyme development during ascidian embryogenesis

Development ◽  
1981 ◽  
Vol 64 (1) ◽  
pp. 61-71
Author(s):  
Noriyuki Satoh ◽  
Susumu Ikegami
Keyword(s):  
Dna Replication ◽  
Nuclear Envelope ◽  
Ache Activity ◽  
Specific Inhibitor ◽  
Specific Enzyme ◽  
Cell Stage ◽  
Tissue Specific ◽  
Nuclear Envelope Breakdown ◽  
Enzyme Development ◽  
Clock Mechanism

Acetylcholinesterase (AChE) is a tissue-specific enzyme of the muscle cells of ascidian embryos and its synthesis begins at the neurula stage. Embryos which had been permanently cleavage-arrested with cytochalasin B could develop AChE activity. The time of first AChE occurrence in embryos which had been arrested in the 32-cell stage with cytochalasin was about the same as in normal embryos. The nucleus in the cell of cytochalasin-arrested embryos divided in good synchrony with that of normal embryos. Embryos which had been continuously arrested with colchicine could also produce AChE activity at nearly the same time as did normal embryos. In the cell of colchicine-arrested embryos normal nuclear divisions did not occur, but the cell showed repeated cycles of nuclear envelope breakdown and nuclear envelope reformation in almost parallel with cell cycles of normal embryos. The cell of colchicine-arrested embryos incorporated [3H]thymidine. Aphidicolin, a specific inhibitor of DNA synthesis, prevented cleavages of ascidian eggs. Embryos which had been permanently arrested with aphidicolin in the cleavage stages up to the 64-cell stage did not develop AChE activity, while embryos which had been treated with it from the 76-cell stage onwards were found to be able to differentiate AChE activity. Based on these findings it was proposed that DNA replication is prerequisite for development of the histospecific protein and that the cycle of DNA replication is closely associated with the clock mechanism which is determining the time of initiation of the enzyme development.

On the ‘clock’ mechanism determining the time of tissue-specific enzyme development during ascidian embryogenesis

Development ◽  
1979 ◽  
Vol 54 (1) ◽  
pp. 131-139
Author(s):  
Noriyuki Satoh
Keyword(s):  
Ache Activity ◽  
Mitotic Cycle ◽  
Cytochalasin B ◽  
Specific Enzyme ◽  
Cleavage Stage ◽  
Halocynthia Roretzi ◽  
Cell Stage ◽  
Tissue Specific ◽  
Enzyme Development ◽  
Clock Mechanism

During ascidian embryogenesis a tissue-specific enzyme, muscle acetylcholinesterase (AChE) may first be detected histochemically in the presumptive muscle cells of the neurula. [n order to investigate the ‘clock’ or counting mechanism that is determining the time when AChE first appears, Whittaker's experiment (1973) has been repeated using eggs of theascidian, Halocynthia roretzi. Embryos that had been permanently cleavage-arrested with cytochalasin B were able to differentiate AChE in their muscle lineage blastomeres. The time of first AChE occurrence in embryos that had been cleavage-arrested in the 32-cell stage with cytochalasin B was about the same as in normal embryos. This result indicates that the clock is not apparently regulated by the events of cytokinesis. The early gastrulae which had been arrested with colchicine or with colcemid could develop AChE activity, although no histochemically detectable AChE activity was observed in the cleavage-stage embryos that had been arrested with either drug. Therefore the clock does not seem to be controlled by the mitotic cycle of the nucleus. It is suggested that the cycle of DNA replication may be related to the regulation of the clock that is determining the time of development of histospecific protein.

Histospecific acetylcholinesterase development in the presumptive muscle cells isolated from 16-cell-stage ascidian embryos with respect to the number of DNA replications

Development ◽  
1985 ◽  
Vol 87 (1) ◽  
pp. 1-12
Author(s):  
Izumi Mita-Miyazawa ◽  
Susumu Ikegami ◽  
Noriyuki Satoh
Keyword(s):  
Dna Replication ◽  
Specific Inhibitor ◽  
Muscle Cells ◽  
Acetyl Cholinesterase ◽  
Specific Enzyme ◽  
Isolated Cells ◽  
Cell Stage ◽  
Simultaneous Treatment ◽  
Tissue Specific ◽  
Ascidian Embryos

The presumptive muscle cells (B5.1 blastomeres) were isolated from 16-cell-stage embryos of the ascidian, Ciona intestinalis. The isolated cells were allowed to divide either twice or three times thereafter. Then further divisions of the cells were continuously inhibited by a simultaneous treatment with aphidicolin (a specific inhibitor of DNA synthesis) and cytochalasin B (an inhibitor of cytokinesis). When development of muscle-specific acetyl-cholinesterase in these division-arrested progeny cells of B5.1 blastomeres was examined histochemically, the B5.1 blastomeres which had been allowed two further divisions did not produce any detectable acetyl-cholinesterase activity. Whereas those which had been allowed three further divisions showed the tissue-specific enzyme activity. These results provide further evidence for the presence of a quantal DNA replication cycle for the tissue-specific enzyme development, which is qualitatively different from the other DNA replication cycles.

A definite number of aphidicolin-sensitive cell-cyclic events are required for acetylcholinesterase development in the presumptive muscle cells of the ascidian embryos

Development ◽  
1981 ◽  
Vol 61 (1) ◽  
pp. 1-13
Author(s):  
Noriyuki Satoh ◽  
Susumu Ikegami
Keyword(s):  
Ache Activity ◽  
Cellular Differentiation ◽  
Cytochalasin B ◽  
Muscle Cells ◽  
Sensitive Cell ◽  
Specific Enzyme ◽  
Cell Stage ◽  
Simultaneous Treatment ◽  
Enzyme Development ◽  
Ascidian Embryos

In order to determine whether or not a crucial number of DNA replications are prerequisite for cellular differentiation, we have studied development of a tissue-specific enzyme, muscle acetylcholinesterase (AChE) in the presumptive muscle cells of cleavage-arrested ascidian embryos. Embryos were cleavage-arrested with cytochalasin B (an inhibitor of cytokinesis) and aphidicolin (an inhibitor of DNA synthesis). The 64-ceIl-stage embryos which had been permanently cleavage-arrested with cytochalasin B developed AChE in all the eight presumptive muscle cells, but the same stage embryos which had been prevented from undergoing further divisions by simultaneous treatment with aphidicolin and cytochalasin did not produce AChE at all. Cytochalasin-arrested 76-cell-stage embryos were able to differentiate AChE in the ten presumptive muscle cells, while aphidicolin-cytochalasin-arrested 76-cell stages in as many as four cells. The early gastrulae which had been arrested with cytochalasin B produced AChE in all the sixteen presumptive muscle cells, while the same stage embryos arrested with aphidicolin and cylochalasin in as many as twelve cells. Cytochalasin-arrested late gastrulae developed AChE in twenty blastomeres, while aphidicolin-cytochalasinarrested late gastrulae in eighteen cells. The presumptive muscle cells at these four stages consist of those of three different (seventh, eighth, and ninth) generations, and the relative positions of the blastomeres in the cleavagearrested embryos remained fixed. Judging from the relative positions of the blaslomeres, the AChE-producing cells in aphidicolin-cytochalasin-arrested embryos were always at eighth or ninth generation, while those with no AChE activity were certainly at seventh generation. Based on these findings it was supposed that aphidicolin-sensitive cell-cyclic events, presumably DNA replication, are closely associated with AChE development and that the eighth cleavage cycle may be ‘quantal’ for the histospecific enzyme development.

scholarly journals Ongoing replication forks delay nuclear envelope breakdown upon mitotic entry

2020 ◽  
pp. jbc.RA120.015142
Author(s):  
Yoshitami Hashimoto ◽  
Hirofumi Tanaka
Keyword(s):  
Dna Replication ◽  
Nuclear Envelope ◽  
Dependent Manner ◽  
Repair Synthesis ◽  
Nuclear Envelope Breakdown ◽  
M Phase ◽  
Egg Extracts ◽  
Fork Stalling ◽  
Dna Repair Synthesis ◽  
Xenopus Egg Extracts

DNA replication is a major contributor to genomic instability and protection against DNA replication perturbation is essential for normal cell division. Certain types of replication stress agents, such as aphidicolin and hydroxyurea, have been shown to cause reversible replication fork stalling, wherein replisome complexes are stably maintained with competence to restart in the S-phase of the cell cycle. If these stalled forks persist into the M-phase without a replication restart, replisomes are disassembled in a p97-dependent pathway and under-replicated DNA is subjected to mitotic DNA repair synthesis. Here, using Xenopus egg extracts, we investigated the consequences that arise when stalled forks are released simultaneously with the induction of mitosis. Ara-cytidine-5’-triphosphate (Ara-CTP)-induced stalled forks were able to restart with the addition of excess dCTPduring early mitosis before the nuclear envelope breakdown (NEB). However, stalled forks could no longer restart efficiently after NEB. Although replisome complexes were finally disassembled in a p97-dependent manner during mitotic progression whether or not fork stalling was relieved, the timing of NEB was delayed with the ongoing forks, rather than the stalled forks, and the delay was dependent on Wee1/Myt1 kinase activities. Thus, ongoing DNA replication was found to be directly linked to the regulation of Wee1/Myt1 kinases to modulate cyclin-dependent kinase (CDK) activities, owing to which DNA replication and mitosis occur in a mutually exclusive and sequential manner.

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