scholarly journals Primase p49 mRNA expression is serum stimulated but does not vary with the cell cycle.

1989 ◽  
Vol 9 (5) ◽  
pp. 1940-1945 ◽  
Author(s):  
B Y Tseng ◽  
C E Prussak ◽  
M T Almazan
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Mammalian Cells ◽  
Cell Cycle Progression ◽  
Small Subunit ◽  
Mrna Levels ◽  
Proliferating Cells ◽  
Restriction Point ◽  
Serum Stimulation ◽  
G1 Cells

Expression of the small-subunit p49 mRNA of primase, the enzyme that synthesizes oligoribonucleotides for initiation of DNA replication, was examined in mouse cells stimulated to proliferate by serum and in growing cells. The level of p49 mRNA increased approximately 10-fold after serum stimulation and preceded synthesis of DNA and histone H3 mRNA by several hours. Expression of p49 mRNA was not sensitive to inhibition by low concentrations of cycloheximide, which suggested that the increase in mRNA occurred before the restriction point control for cell cycle progression described for mammalian cells and was not under its control. p49 mRNA levels were not coupled to DNA synthesis, as observed for the replication-dependent histone genes, since hydroxyurea or aphidicolin had no effect on p49 mRNA levels when added before or during S phase. These inhibitors did have an effect, however, on the stability of p49 mRNA and increased the half-life from 3.5 h to about 20 h, which suggested an interdependence of p49 mRNA degradation and DNA synthesis. When growing cells were examined after separation by centrifugal elutriation, little difference was detected for p49 mRNA levels in different phases of the cell cycle. This was also observed when elutriated G1 cells were allowed to continue growth and then were blocked in M phase with colcemid. Only a small decrease in p49 mRNA occurred, whereas H3 mRNA rapidly decreased, when cells entered G2/M. These results indicate that the level of primase p49 mRNA is not cell cycle regulated but is present constitutively in proliferating cells.

Primase p49 mRNA expression is serum stimulated but does not vary with the cell cycle

1989 ◽  
Vol 9 (5) ◽  
pp. 1940-1945
Author(s):  
B Y Tseng ◽  
C E Prussak ◽  
M T Almazan
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Mammalian Cells ◽  
Cell Cycle Progression ◽  
Small Subunit ◽  
Mrna Levels ◽  
Proliferating Cells ◽  
Restriction Point ◽  
Serum Stimulation ◽  
G1 Cells

Expression of the small-subunit p49 mRNA of primase, the enzyme that synthesizes oligoribonucleotides for initiation of DNA replication, was examined in mouse cells stimulated to proliferate by serum and in growing cells. The level of p49 mRNA increased approximately 10-fold after serum stimulation and preceded synthesis of DNA and histone H3 mRNA by several hours. Expression of p49 mRNA was not sensitive to inhibition by low concentrations of cycloheximide, which suggested that the increase in mRNA occurred before the restriction point control for cell cycle progression described for mammalian cells and was not under its control. p49 mRNA levels were not coupled to DNA synthesis, as observed for the replication-dependent histone genes, since hydroxyurea or aphidicolin had no effect on p49 mRNA levels when added before or during S phase. These inhibitors did have an effect, however, on the stability of p49 mRNA and increased the half-life from 3.5 h to about 20 h, which suggested an interdependence of p49 mRNA degradation and DNA synthesis. When growing cells were examined after separation by centrifugal elutriation, little difference was detected for p49 mRNA levels in different phases of the cell cycle. This was also observed when elutriated G1 cells were allowed to continue growth and then were blocked in M phase with colcemid. Only a small decrease in p49 mRNA occurred, whereas H3 mRNA rapidly decreased, when cells entered G2/M. These results indicate that the level of primase p49 mRNA is not cell cycle regulated but is present constitutively in proliferating cells.

Microinjection of fos-specific antibodies blocks DNA synthesis in fibroblast cells

1988 ◽  
Vol 8 (4) ◽  
pp. 1670-1676 ◽  
Author(s):  
K T Riabowol ◽  
R J Vosatka ◽  
E B Ziff ◽  
N J Lamb ◽  
J R Feramisco
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Antisense Rna ◽  
Proliferating Cells ◽  
Rna Transcripts ◽  
Serum Stimulation ◽  
Inhibition Of Growth ◽  
Nonspecific Immunoglobulins ◽  
3T3 Cell Lines ◽  
Stimulation Of

Transcription of the protooncogene c-fos is increased greater than 10-fold within minutes of treatment of fibroblasts with serum or purified growth factors. Recent experiments with mouse 3T3 cell lines containing inducible fos antisense RNA constructs have shown that induced fos antisense RNA transcripts cause either a marked inhibition of growth in continuously proliferating cells or, conversely, a minimal effect except during the transition from a quiescent (G0) state into the cell cycle. Since intracellular production of large amounts of antisense RNA does not completely block gene expression, we microinjected affinity-purified antibodies raised against fos to determine whether and when during the cell cycle c-fos expression was required for cell proliferation. Using this independent method, we found that microinjected fos antibodies efficiently blocked serum-stimulated DNA synthesis when injected up to 6 to 8 h after serum stimulation of quiescent REF-52 fibroblasts. Furthermore, when fos antibodies were injected into asynchronously growing cells, a consistently greater number of cells was prevented from synthesizing DNA than when cells were injected with nonspecific immunoglobulins. Thus, whereas the activity of c-fos may be necessary for transition of fibroblasts from G0 to G1 of the cell cycle, its function is also required during the early G1 portion of the cell cycle to allow subsequent DNA synthesis.

scholarly journals A comprehensive model for the proliferation–quiescence decision in response to endogenous DNA damage in human cells

2018 ◽  
Vol 115 (10) ◽  
pp. 2532-2537 ◽  
Author(s):  
Frank S. Heldt ◽  
Alexis R. Barr ◽  
Sam Cooper ◽  
Chris Bakal ◽  
Béla Novák
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Kinase Inhibitor ◽  
Human Cells ◽  
Cyclin Dependent Kinase ◽  
Restriction Point ◽  
Cyclin Dependent Kinase Inhibitor ◽  
Serum Stimulation ◽  
G1 Cells ◽  
External Stimuli

Human cells that suffer mild DNA damage can enter a reversible state of growth arrest known as quiescence. This decision to temporarily exit the cell cycle is essential to prevent the propagation of mutations, and most cancer cells harbor defects in the underlying control system. Here we present a mechanistic mathematical model to study the proliferation–quiescence decision in nontransformed human cells. We show that two bistable switches, the restriction point (RP) and the G1/S transition, mediate this decision by integrating DNA damage and mitogen signals. In particular, our data suggest that the cyclin-dependent kinase inhibitor p21 (Cip1/Waf1), which is expressed in response to DNA damage, promotes quiescence by blocking positive feedback loops that facilitate G1 progression downstream of serum stimulation. Intriguingly, cells exploit bistability in the RP to convert graded p21 and mitogen signals into an all-or-nothing cell-cycle response. The same mechanism creates a window of opportunity where G1 cells that have passed the RP can revert to quiescence if exposed to DNA damage. We present experimental evidence that cells gradually lose this ability to revert to quiescence as they progress through G1 and that the onset of rapid p21 degradation at the G1/S transition prevents this response altogether, insulating S phase from mild, endogenous DNA damage. Thus, two bistable switches conspire in the early cell cycle to provide both sensitivity and robustness to external stimuli.

scholarly journals The jun and fos protein families are both required for cell cycle progression in fibroblasts.

1991 ◽  
Vol 11 (9) ◽  
pp. 4466-4472 ◽  
Author(s):  
K Kovary ◽  
R Bravo
Keyword(s):  
Cell Cycle ◽  
Transcription Factors ◽  
Dna Synthesis ◽  
Cell Cycle Progression ◽  
S Phase ◽  
3T3 Cells ◽  
Cycle Progression ◽  
Serum Stimulation ◽  
Fos Protein ◽  
Stimulation Of

The expression of different members of the Jun and Fos families of transcription factors is rapidly induced following serum stimulation of quiescent fibroblasts. To determine whether these proteins are required for cell cycle progression, we microinjected affinity-purified antibodies directed against c-Fos, FosB, Fra-1, c-Jun, JunB, and JunD, and antibodies that recognize either the Fos or the Jun family of proteins, into Swiss 3T3 cells and determined their effects in cell cycle progression by monitoring DNA synthesis. We found that microinjection of anti-Fos and anti-Jun family antibodies efficiently blocked the entrance to the S phase of serum-stimulated or asynchronously growing cells. However, the antibodies against single members of the Fos family only partially inhibited DNA synthesis. In contrast, all three Jun antibodies prevented DNA synthesis more effectively than did any of the anti-Fos antibodies.

scholarly journals Microinjection of fos-specific antibodies blocks DNA synthesis in fibroblast cells.

1988 ◽  
Vol 8 (4) ◽  
pp. 1670-1676 ◽  
Author(s):  
K T Riabowol ◽  
R J Vosatka ◽  
E B Ziff ◽  
N J Lamb ◽  
J R Feramisco
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Antisense Rna ◽  
Proliferating Cells ◽  
Rna Transcripts ◽  
Serum Stimulation ◽  
Inhibition Of Growth ◽  
Nonspecific Immunoglobulins ◽  
3T3 Cell Lines ◽  
Stimulation Of

Transcription of the protooncogene c-fos is increased greater than 10-fold within minutes of treatment of fibroblasts with serum or purified growth factors. Recent experiments with mouse 3T3 cell lines containing inducible fos antisense RNA constructs have shown that induced fos antisense RNA transcripts cause either a marked inhibition of growth in continuously proliferating cells or, conversely, a minimal effect except during the transition from a quiescent (G0) state into the cell cycle. Since intracellular production of large amounts of antisense RNA does not completely block gene expression, we microinjected affinity-purified antibodies raised against fos to determine whether and when during the cell cycle c-fos expression was required for cell proliferation. Using this independent method, we found that microinjected fos antibodies efficiently blocked serum-stimulated DNA synthesis when injected up to 6 to 8 h after serum stimulation of quiescent REF-52 fibroblasts. Furthermore, when fos antibodies were injected into asynchronously growing cells, a consistently greater number of cells was prevented from synthesizing DNA than when cells were injected with nonspecific immunoglobulins. Thus, whereas the activity of c-fos may be necessary for transition of fibroblasts from G0 to G1 of the cell cycle, its function is also required during the early G1 portion of the cell cycle to allow subsequent DNA synthesis.

Dolichol delays G1-arrest for one cell cycle in human fibroblasts subjected to depletion of serum or mevalonate

1992 ◽  
Vol 103 (4) ◽  
pp. 1065-1072 ◽  
Author(s):  
O. Larsson ◽  
J. Wejde
Keyword(s):  
Cell Cycle ◽  
Mammalian Cells ◽  
Cell Cycle Progression ◽  
Cell Cycle Control ◽  
Human Fibroblasts ◽  
Video Recording ◽  
Time Lapse ◽  
G1 Arrest ◽  
Proliferating Cells ◽  
Serum Factors

It is well-established that some product(s) or metabolite(s) of mevalonate is (are) critical for growth of mammalian cells. In the search for this (these) compound(s) it seems meaningful to distinguish between compounds needed for cell cycle progression in proliferating cells and compounds needed for growth activation of arrested cells. By using time-lapse video recording we have studied the possible regulatory role of cholesterol, dolichol and mevalonate in the cell cycle of human diploid fibroblasts (HDF). HDF, which are serum-dependent, were rapidly growth-arrested in the first part of G1 upon removal of serum factors. They also responded to mevinolin (an HMG CoA reductase inhibitor) by a similar G1-block, indicating that a mevalonate-derived product is involved in the G1-located cell cycle control of HDF. Interestingly, dolichol counteracted the G1-block caused by both types of treatment. Hence, the early G1-cells could traverse the remainder of the cell cycle and divide despite depletion of serum or mevalonate. We also demonstrated that addition of dolichol resulted in a significant decrease in the rate of protein degradation. This protein stabilizing effect may constitute the mechanism by which dolichol delays the G1-arrest of HDF.

Periodic expression of nuclear and mitochondrial DNA replication genes during the trypanosomatid cell cycle

1994 ◽  
Vol 107 (12) ◽  
pp. 3515-3520
Author(s):  
S.G. Pasion ◽  
G.W. Brown ◽  
L.M. Brown ◽  
D.S. Ray
Keyword(s):  
Cell Cycle ◽  
Mitochondrial Dna ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Cell Cycle Progression ◽  
Nuclear Dna ◽  
Protein A ◽  
Mrna Levels ◽  
Gene Encoding ◽  
Genes Encoding

In trypanosomatids, DNA replication in the nucleus and in the single mitochondrion (or kinetoplast) initiates nearly simultaneously, suggesting that the DNA synthesis (S) phases of the nucleus and the mitochondrion are coordinately regulated. To investigate the basis for the temporal link between nuclear and mitochondrial DNA synthesis phases the expression of the genes encoding DNA ligase I, the 51 and 28 kDa subunits of replication protein A, dihydrofolate reductase and the mitochondrial type II topoisomerase were analyzed during the cell cycle progression of synchronous cultures of Crithidia fasciculata. These DNA replication genes were all expressed periodically, with peak mRNA levels occurring just prior to or at the peak of DNA synthesis in the synchronized cultures. A plasmid clone (pdN-1) in which TOP2, the gene encoding the mitochondrial topoisomerase, was disrupted by the insertion of a NEO drug-resistance cassette was found to express both a truncated TOP2 mRNA and a truncated topoisomerase polypeptide. The truncated mRNA was also expressed periodically coordinate with the expression of the endogenous TOP2 mRNA indicating that cis elements necessary for periodic expression are contained within cloned sequences. The expression of both TOP2 and nuclear DNA replication genes at the G1/S boundary suggests that regulated expression of these genes may play a role in coordinating nuclear and mitochondrial S phases in trypanosomatids.

scholarly journals Chamber-specific differences in human cardiac fibroblast proliferation and responsiveness toward simvastatin

2016 ◽  
Vol 311 (2) ◽  
pp. C330-C339 ◽  
Author(s):  
Farhan Rizvi ◽  
Alessandra DeFranco ◽  
Ramail Siddiqui ◽  
Ulugbek Negmadjanov ◽  
Larisa Emelyanova ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Cell Cycle Progression ◽  
Cardiac Fibrosis ◽  
Ventricular Remodeling ◽  
Cardiac Fibroblasts ◽  
Mrna Levels ◽  
Cycle Progression ◽  
Hmg Coa Reductase ◽  
Coa Reductase

Fibroblasts, the most abundant cells in the heart, contribute to cardiac fibrosis, the substrate for the development of arrythmogenesis, and therefore are potential targets for preventing arrhythmic cardiac remodeling. A chamber-specific difference in the responsiveness of fibroblasts from the atria and ventricles toward cytokine and growth factors has been described in animal models, but it is unclear whether similar differences exist in human cardiac fibroblasts (HCFs) and whether drugs affect their proliferation differentially. Using cardiac fibroblasts from humans, differences between atrial and ventricular fibroblasts in serum-induced proliferation, DNA synthesis, cell cycle progression, cyclin gene expression, and their inhibition by simvastatin were determined. The serum-induced proliferation rate of human atrial fibroblasts was more than threefold greater than ventricular fibroblasts with faster DNA synthesis and higher mRNA levels of cyclin genes. Simvastatin predominantly decreased the rate of proliferation of atrial fibroblasts, with inhibition of cell cycle progression and an increase in the G0/G1 phase in atrial fibroblasts with a higher sensitivity toward inhibition compared with ventricular fibroblasts. The DNA synthesis and mRNA levels of cyclin A, D, and E were significantly reduced by simvastatin in atrial but not in ventricular fibroblasts. The inhibitory effect of simvastatin on atrial fibroblasts was abrogated by mevalonic acid (500 μM) that bypasses 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibition. Chamber-specific differences exist in the human heart because atrial fibroblasts have a higher proliferative capacity and are more sensitive to simvastatin-mediated inhibition through HMG-CoA reductase pathway. This mechanism may be useful in selectively preventing excessive atrial fibrosis without inhibiting adaptive ventricular remodeling during cardiac injury.

Functional conservation of the cell cycle-regulating transcription factor DRTF1/E2F and its pathway of control in Drosophila melanogaster

1995 ◽  
Vol 108 (9) ◽  
pp. 2945-2954 ◽  
Author(s):  
X.F. Hao ◽  
L. Alphey ◽  
L.R. Bandara ◽  
E.W. Lam ◽  
D. Glover ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Transcription Factor ◽  
Dna Binding ◽  
Mammalian Cells ◽  
Cell Cycle Progression ◽  
Cellular Proliferation ◽  
Expression Patterns ◽  
Proliferating Cells ◽  
Cycle Progression ◽  
Pocket Proteins

The cellular transcription factor DRTF1/E2F is implicated in the control of early cell cycle progression due to its interaction with important regulators of cellular proliferation, such as pocket proteins (for example, the retinoblastoma tumour suppressor gene product), cyclins and cyclin-dependent kinase subunits. In mammalian cells DRTF1/E2F is a heterodimeric DNA binding activity which arises when a DP protein interacts with an E2F protein. Here, we report an analysis of DRTF1/E2F in Drosophila cells, and show that many features of the pathway which regulate its transcriptional activity are conserved in mammalian cells, such as the interaction with pocket proteins, binding to cyclin A and cdk2, and its modulation by viral oncoproteins. We show that a Drosophila DP protein which can interact co-operatively with E2F proteins is a physiological DNA binding component of Drosophila DRTF1/E2F. An analysis of the expression patterns of a Drosophila DP and E2F protein indicated that DmDP is developmentally regulated and in later embryonic stages preferentially expressed in proliferating cells. In contrast, the expression of DmE2F-1 in late stage embryos occurs in a restricted group of neural cells, whereas in early embryos it is widely expressed, but in a segmentally restricted fashion. Some aspects of the mechanisms which integrate early cell cycle progression with the transcription apparatus are thus conserved between Drosophila and mammalian cells. The distinct expression patterns of DmDP and DmE2F-1 suggest that the formation of DP/E2F heterodimers, and hence DRTF1/E2F, is subject to complex regulatory cues.

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