Arrest of DNA synthesis

As embryogenesis involves extensive cellular proliferation, any interference with this process is potentially teratogenic. Interference with spindle formation, inhibition or arrest of DNA synthesis or the incorrect separation of chromatids all fall into this category. Inhibition of DNA synthesis, such as that caused by cytosine arabinoside, slows or arrests mitosis, which cannot progress beyond the S-phase. Thus those areas of the embryo which show extensive cellular proliferation are the most susceptible to both necrosis and subsequent malformation from cytotoxic compounds such as cytosine arabinoside. Inhibition of spindle formation such as that caused by vincristine or colchicine stops separation of chromosomes at anaphase (see page 240). Proper separation of chromatids may not occur because of stickiness or bridging between the chromatids. Clearly interference with mitosis, and hence cell proliferation, is an important cause of teratogenic effects. 

Folic acid antagonist inhibits dihydrofolate reductase (DHFR) blocks reduction of folate to tetrahydrofolate inhibits de novo purine synthesis results in arrest of DNA, RNA, and protein synthesis... 

BrdU analysis allows to distinguish and quantify if there is an arrest in DNA synthesis, in which compartment of S phase there are and it is possible to separate E (early) S from Gi or L (Late) S from G2/M. This advantage is demonstrated in Figure 5, where HCT-116 cells were treated with SN-38 or edotecarin that affect DNA replication [21],... 

Proposed mechanism of action of daptomycin. Daptomycin first binds to the cytoplasmic membrane (step 1) and then forms complexes in a calcium-dependent manner (steps 2 and 3). Complex formation causes a rapid loss of cellular potassium, possibly by pore formation, and membrane depolarization. This is followed by arrest of DNA, RNA, and protein synthesis resulting in cell death. Cell lysis does not occur. 

Temin (1970) and Todaro et al. (1965) showed similar effects for chicken fibroblasts and 3T3 mouse fibroblasts. The low level of serum is important for survival as well as for the subsequent stimulation of DNA synthesis (Cherrington, 1984). A kinetic analysis using time lapse cinematography (Zetterberg and Larsson, 1985) showed that Swiss 3T3 cells were only susceptible to cell cycle arrest in a short period (3-4 h) following mitosis. Even a 1-h exposure to serum-free medium during this time forced the cells into GO from which they required 8 h to return to Gl. The length of the postmitotic sensitive phase was very constant at between 3 and 4 h but considerable intercellular variability existed in the duration of the pre S-phase Gl period consistent with a transition probability event ( 10.4). 

Cells treated with 5-FU undergo cell cycle arrest or apoptosis by inhibition of DNA synthesis. When human oral squamous cell carcinomas (HSC-2, HSC-3, HSC-4) were treated with 5-FU, viable cell numbers declined dose-dependently (CC50 determined after 24 hours treatment 3.4,6.9 and 1.7 p,M, respectively). The intracellular putrescine concentration slightly declined (Table 2) [29]. The combination treatment of 5-FU with N(l),N(ll)-diethylnorspermine (DEN-SPM), which is an inducer of spermidine/spermine N(l)-acetyltransferase (SSAT) that depletes polyamine, has been reported to augment the cytotoxic activity of 5-FU, suggesting possible clinical application [30]. 

Holotoxin Ai inhibits the RNA biosynthesis in Candida albicans and Saccharomyces carlsbergensis, as indicated by the decrease in incorporation of C-uridine to the acid-insoluble fraction of the cells. Similar results were obtained for glycoside fractions of 14 species of Pacific sea cucumbers [132]. Apparently the inhibition of RNA biosynthesis in Saccharomyces carlsbergensis is related to nucleotide loss from yeast cells after treatment with glycosides. Holotoxin Ai also inhibits biosyntheses of squalene, lanosterol and ergosterol in S. carlsbergensis [133]. Mitosis is arrested and DNA synthesis inhibited in onion root bulbs by crude holothurin [134]. 

Azacytidine, after conversion to a phosphorylated derivative, arrests the DNA synthesis phase of the cell cycle. It also affects methylation of certain bases, which leads to hypomethylation (Chapter 26). 

Nocodazole, a drug that inhibits the assembly of microtubules, arrests normal mouse embryo fibroblasts with a 4N content, whereas fibroblasts from p53-deficient mouse embryos undergo several rounds of DNA synthesis without chromosome segregation (Cross et al., 1995). This spindle checkpoint differs from the G2/M checkpoint. Both p53+/+ and p53 / fibroblasts treated with nocodazole transiently arrest at mitosis for the same length of... 

The 5-bromo and 5-chloro derivatives of l-propargyl-2(l//)-pyrimidinones are inhibitors of the micotubule system in malignant cells. In rat glioma cells, arrest of mitosis was accompanied by repeated cycles of DNA synthesis, leading to different levels of polyploidization with formation of up to 16 and 32 ploid cells <86MI 602-04>. 

The growth inhibition of virus-transformed cells in vitro and antitumor activity in vivo of geldanamycin (85) and its derivatives [230,231], and the inhibition of DNA synthesis in murine tumor cells by 85 were reported [232,233]. Geldanamycin (85) is an antibiotic that preferentially inhibits Gl/S transition and causes G2/M arrest in human leukemia HL-60 cells. Also, it was found that 85 selectively inhibited recombinant Src tyrosine kinase without significantly inhibiting protein kinase A. The perturbation of cell cycling by 85 was accompanied by marked suppression of c-MYC expression [234]. 

The role of oxygen in eukaryotic DNA biosynthesis may indeed be a critical one. It has recently been shown that O2 is not only required for initial formation of tyrosyl radical but must be continously present to maintain the radical content and enzyme activity of mammalian ribonucleotide reductase In vivo studies with Ehrlich ascites cells also point to a tight link between oxygen and deoxyribonucleotide supply Anaerobic arrest of cells in G1 phase and block of DNA synthesis can be relieved by addition of deoxycytidine, but not cytidine, to the culture medium. ... 

The initiation or arrest of enzyme synthesis responsible for the peak of ribonucleotide reductase is discussed below. Other factors which may modulate the activity in vivo or when measured in crude homogenates include allosteric control by the endogenous deoxyribonucleotides, the action of late S phase-specific inhibitors like the one found in Achlya, or the redox status of thioredoxin and glutaredoxin however the cell cycle dependence of these reactions is little known. It is therefore desirable to assay ribonucleotide reductase in preparations which have been subjected to at least one precipitation and dialysis step. While reliable cell cycle-dependent activity data are thus obtained, the absolute figures are frequently an order of magnitude too low to account for the cell s need of ribonucleotide reduction for DNA synthesis. This unsatisfactory condition is most likely a problem of quinary enzyme structure (see below) but is not felt to invalidate the accumulated evidence for tight correlation of ribonucleotide reduction and the cell cycle as a whole. 

Daptomycin was originally isolated from the soil saprotroph Streptomyces roseos-porus by scientists at Eli Lilly and Company in the 1980s. It is a novel lipopeptide antibiotic used in the treatment of certain infections caused by Gram-positive organisms. The proposed mechanism of action involves insertion of the lipophilic daptomycin tail into the bacterial cell membrane, causing rapid membrane depolarization and a potassium ion efflux. This leads to the arrest of DNA, RNA, and protein synthesis, resulting in bacterial cell death (see footnote 1). 

No events have been described which uniquely characterize the G1 phase. The G1 phase appears to perform two functions the temporary or permanent arrest of cell division in eukaryotes is normally achieved by cells maintaining themselves in the G1 phase of the cell cycle (Prescott, 1970) also the terminal part of the G1 must contain events concerned with the initiation of DNA synthesis. Certain organisms, for example, the fission yeast Schizosaccharomyces pombe (Bostock et al., 1966), the multinucleate slime mold Physarum polycephalum (Guttes et al., 1967), and Amoeba proteus (Ord, 1968), do not exhibit a G1 phase. A number of mammaUan cells appear to lack a G1... 

The contribution of the GV to either the induction of chromosome condensation in transplanted nuclei or the induction of DNA synthesis is not yet clear. Masui and Markert (1971) have reported that cytoplasm from maturing Rana pipiens oocytes inhibits cell division when injected into fertilized eggs or cleavage blastomeres. One effect of this cytostatic factor is to cause mitosis to arrest at metaphase. The cytostatic factor appears only after breakdown of the GV, but this temporal coincidence is misleading enucleated oocytes acquire the factor at about the same time (Masui and Markert, 1971). Thus, to the extent that the cytostatic factor is the same as that which leads to chromosomal condensation and attachment to spindles in transplanted nuclei (Gurdon, 1968), neither the GV nor its contents makes a direct contribution to the appearance of such material. It is not known whether oocytes, enucleated prior to hormone exposure, subsequently acquire the cytoplasmic capacity to induce DNA synthesis in transplanted nuclei. 

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