Growth-dependent synthesis of c-myc-encoded proteins: early stimulation by serum factors in synchronized mouse 3T3 cells.

Persson, H; Gray, H E; Godeau, F

doi:10.1128/mcb.5.11.2903

Cited by 53 publications

(49 citation statements)

References 40 publications

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“…Mitosis occurred approximately 27 h after serum stimulation. These data are similar to those obtained by others (3,24). To assess drug sensitivity throughout the cell cycle, cells were exposed to etoposide for 1 h at various times after serum stimulation and DNA cleavage was assayed by the alkaline elution technique.…”

supporting

confidence: 64%

Topoisomerase-specific drug sensitivity in relation to cell cycle progression.

Chow¹,

Ross²

1987

Mol. Cell. Biol.

134

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. It also mediates the DNA cleavage activity and cytotoxicity of clinically important anticancer agents such as etoposide. We have examined the activity of topoisomerase H during the first cell cycle of quiescent BALB/c 3T3 cells following serum stimulation. Etoposide-mediated DNA break frequency in vivo was used as a parameter of topoisomerase II activity, and enzyme content was assayed by immunoblotting. Density-arrested A31 cells exhibited a much lower sensitivity to the effects of etoposide than did actively proliferating cells. Upon serum stimulation of the quiescent cells, however, there was a marked increase in drug sensitivity which began during S phase and reached its peak just before mitosis. Maximal drug sensitivity during this period was 2.5 times greater than that of log-phase cells. This increase in drug sensitivity was associated with an increase in intracellular topoisomerase II content as determined by immunoblotting. The induction of topoisomerase 11-mediated drug sensitivity was aborted within 1 h of exposure of cells to the protein synthesis inhibitor cycloheximide, but the DNA synthesis inhibitor aphidicolin had no effect. In contrast to the sensitivity of cells to drug-induced DNA cleavage, maximal cytotoxicity occurred during S phase. A 3-h exposure to cycloheximide before etoposide treatment resulted in nearly complete loss of cytotoxicity. Our findings indicate that topoisomerase II activity fluctuates with cell cycle progression, with peak activity occurring during the G2 phase. This increase in topoisomerase II is protein synthesis dependent and may reflect a high rate of enzyme turnover. The dissociation between maximal drug-induced DNA cleavage and cytotoxicity indicates that the topoisomerase-mediated DNA breaks may be necessary but are not sufficient for cytotoxicity and that other factors which are particularly expressed during S phase may be important as well.Type II DNA topoisomerases are ATP-dependent enzymes which catalyze the breakage and reunion of duplex DNA, allowing a second segment of DNA to pass through the break site (1, 23). They are required for the segregation of DNA following the conclusion of replication (4) and appear to be required at mitosis (12). During the breakagereunion cycle a covalent phosphotyrosyl bond between enzyme and DNA is formed at the 5' terminus of each strand. A number of commonly used anticancer agents such as intercalating agents and epipodophyllotoxins can stabilize this DNA protein intermediate, and convincing evidence exists that this interaction does mediate their cytotoxicity (15,16,25). A better understanding of the enzyme would be useful in improving the efficacy of these drugs. In addition, these topoisomerase-specific drugs have become useful probes in understanding the fundamental functions and control mechanisms of the enzyme.Topoisomerase II activity and content varies as a function of the proliferative status of the cell (5,19,29,30 Cell culture. BALB/c 3T3 (A31) were grown at 37°C in a monolayer in alpha minimal essential medium...

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supporting

confidence: 64%

Topoisomerase-specific drug sensitivity in relation to cell cycle progression.

Chow¹,

Ross²

1987

Mol. Cell. Biol.

134

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“…It has also been shown that myc RNA and protein do not correlate during dierentiation of murine erythroleukaemia cells (Wingrove et al, 1988;Spotts and Hann, 1990). Although most studies have demonstrated that myc transcripts and protein levels are directly related (Hann and Eisenman, 1984;Persson et al, 1985;Rabbitts et al, 1985;Hann et al, 1985), the data presented here support previous reports that translational and posttranslational regulatory mechanisms play an important role in ultimately determining Myc protein content of cells (Darveau et al, 1985;Parkin et al, 1988;Lazarus et al, 1988;Spotts and Hann, 1990). One possible explanation for the unexpectedly low Myc protein content in Em-myc B cell lines involves the ratio of transcripts generated from the two promoters, P 1 and P 2 .…”

Section: Eµ-mycsupporting

confidence: 80%

“…Most previous studies have shown that Myc protein levels correlate well with the RNA content of cells (Hann and Eisenman, 1984;Persson et al, 1985;Rabbitts et al, 1985;Hann et al, 1985). Immunoblotting was used to determine whether the elevated myc mRNA observed in the Em-myc B lymphoma cells Figure 4a show that unlike the considerable dierences in transcript levels (Figure 3), the Myc protein content of Em-myc B cell lines was only 30 ± 50% greater than that of the v-raf derived macrophages.…”

Section: Eµ-mycmentioning

confidence: 99%

“…To investigate whether changes to Myc protein stability may account for the discrepancy between RNA and protein levels in Em-myc cell lines, the turnover rate of the protein was assessed. Myc protein normally has a half life of between 20 and 30 min (Hann and Eisenman, 1984;Rabbitts et al, 1985;Persson et al, 1985). However, alterations to this turnover rate have been observed which can aect accumulation of the protein (Wingrove et al, 1988;Spotts and Hann, 1990).…”

Section: Eµ-mycmentioning

confidence: 99%

“…Finally, the Myc protein content was assessed and compared with the relative abundance of myc RNA. As most investigations have shown a direct relationship between myc RNA and protein (Hann and Eisenman, 1984;Persson et al, 1985;Rabbitts et al, 1985;Hann et al, 1985), it was anticipated that a marked increase in Myc protein would be detected in Em-myc B lymphoma cells that matched the raised RNA levels.…”

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Increased transcription of the Eμ-myc transgene and mRNA stabilisation produce only a modest elevation in Myc protein

1997

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Mice bearing the Em-myc transgene, which links the immunoglobulin heavy chain enhancer (Em) with c-myc, are predisposed to developing B cell lymphomas. Several B lineage cell lines have been isolated from these animals, and some have been converted to macrophages following infection with v-raf. In this study we compared the regulation of myc expression in Em-myc B lymphoma lines, their macrophage counterparts and other non-myc transformed B cell lines. Nuclear run-on analyses demonstrated that transcription of the transgene was elevated in Em-myc B cell lines. Moreover, the presence of a 600 bp fX174 marker in the 3' end of the transgene produced a marked stabilisation of this RNA species. Consequently, steady state myc mRNA levels in the Emmyc B lymphoma cells were tenfold higher than the macrophage derivatives and non-myc transformed B lineage lines. Despite the considerable dierence in myc RNA levels, the Em-myc B cell lines contained only 30 ± 50% more Myc protein than the other cell lines. This discrepancy between RNA and protein content was not due to increased degradation of the protein as the half life was normal in the transgenic cell lines. These results indicate that both Em and fX174 sequences in¯uence transgenic myc expression and that protein levels do not correlate with RNA content in Em-myc cell lines.Keywords: Em-myc transgene; mRNA stabilisation; transcription; Myc protein Expression of the myc gene is subject to an elaborate array of transcriptional, post-transcriptional and translational regulatory mechanisms and perturbation of these controls has been implicated in the genesis of a wide range of tumours (Spencer and Groudine, 1991). Translocations involving immunoglobulin loci and cmyc have been identi®ed in Burkitt's lymphoma and murine plasmacytomas (Cory, 1986). To investigate the role of the immunoglobulin heavy chain enhancer (Em) in these tumours, Adams et al. (1985) generated transgenic mice coupling this enhancer to c-myc. In addition, a fragment of fX174 was introduced into the 3' untranslated region (3'UTR) of the transgene to generate a larger transcript which could easily be distinguished from the endogenous myc mRNA. Expression of Em-myc was restricted to the B lymphoid compartment and transgenic animals contained abnormally high levels of early B cells before succumbing to B lineage lymphomas Langdon et al., 1986;Langdon et al., 1988). Transgene RNA levels were ®ve to sixfold greater in Em-myc splenocytes and cell lines than c-myc in the spleens of non-transgenic littermates (Alexander et al., 1987). It was presumed that`the malignancy is a consequence of high constitutive myc expression within the lymphoid compartment, forced by enhancers which are activated by lymphoid factors . et al., 1985). Labelled transcripts were hybridised to membranes containing probes for myc (Stanton et al., 1983), Cm (Gough et al., 1980) and GAPDH (Piechaczyk et al., 1984). Annealed transcripts were detected by phosphorimager (a) and their levels expressed relative to GAPDH (b). Black histograms ...

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Regulation and Function of the MYC Oncogene

Kalkat

Penn

2013

Encyclopedia of Life Sciences

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The MYC oncogene has been the subject of intense study for more than 30 years, due to its important role in tumourigenesis. MYC is a nuclear transcription factor that can regulate the expression of a multitude of target genes, and its expression in tumours is often associated with poor prognosis. Although decades of research have provided insights into its molecular function as a transcription factor, the mechanisms by which MYC can induce cellular transformation have remained elusive. In addition to the regulation of genes important for cell cycle progression, metabolism, apoptosis and angiogenesis, MYC has wide‐reaching effects, including the alteration of cellular epigenetics and total ribonucleic acid (RNA) content. Recent reports have suggested that post‐translational modification (PTM) of MYC is important for regulating the activity and functions of this potent oncogene; however, surprisingly, little research has been conducted in this area. Herein, the authors summarise recent findings regarding the molecular and biological functions of MYC and their relationship to the PTMs of MYC. Key Concepts: The MYC oncogene encodes a nuclear basic helix‐loop‐helix transcription factor. MYC regulates a large number of transcriptional targets and can have wide‐reaching effects on proliferation, the epigenome and total cellular RNA content. MYC is essential for cellular proliferation and is a potent oncogene when it is deregulated in cancer. Deregulated, often elevated, MYC levels have been shown to initiate tumourigenesis in many in vivo models. MYC activity and stability is regulated by post‐translational modifications, including phosphorylation, ubiquitylation and acetylation.

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Growth-dependent synthesis of c-myc-encoded proteins: early stimulation by serum factors in synchronized mouse 3T3 cells.

Cited by 53 publications

References 40 publications

Topoisomerase-specific drug sensitivity in relation to cell cycle progression.

Topoisomerase-specific drug sensitivity in relation to cell cycle progression.

Increased transcription of the Eμ-myc transgene and mRNA stabilisation produce only a modest elevation in Myc protein

Regulation and Function of the MYC Oncogene

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