scholarly journals DNA-Dependent DNA Polymerase from Yeast Mitochondria. Dependence of Enzyme Activity on conditions of Cell Growth, and Properties of the Highly Purified Polymerase

1976 ◽  
Vol 68 (1) ◽  
pp. 199-207 ◽  
Author(s):  
Ulrike WINTERSBERGER ◽  
Hans BLUTSCH
Keyword(s):  
Enzyme Activity ◽  
Cell Growth ◽  
Dna Polymerase ◽  
Yeast Mitochondria

GENERATIVE CYCLES OF PROTEIN AND TRANSAMINASE IN THE GROWING CORN RADICLE

1959 ◽  
Vol 37 (4) ◽  
pp. 621-639 ◽  
Author(s):  
F. S. Cook
Keyword(s):  
Enzyme Activity ◽  
Cell Growth ◽  
Growth And Development ◽  
Specific Protein ◽  
Soluble Nitrogen ◽  
Transaminase Activity ◽  
Cell Numbers ◽  
Nitrogen Levels ◽  
Linear Distance ◽  
Cellular Development

Measurements have been made of growth rates, cell numbers, fresh and dry weights, protein and soluble nitrogen levels, and glutamic–aspartic transaminase activity in six successive 2 mm segments of the radicles of 3-day-old corn seedlings. The measured quantities of protein and enzyme activity are related to the stage of average cellular development, to a linear distance scale along the axis of the radicle, and to the time scale. Increments per cell per hour during cell growth are therefore computed. An attempt is made to explain the significance of the genesis of the transaminase in the growth and development of the radicle cells, to the concurrent genesis of total and specific protein, and to other generative cycles.

Subcellular Distribution of DNA Polymerase Activity in Newborn Rat Brain and Liver

1971 ◽  
Vol 49 (12) ◽  
pp. 1285-1291 ◽  
Author(s):  
M. R. V. Murthy ◽  
A. D. Bharucha
Keyword(s):  
Enzyme Activity ◽  
Rat Brain ◽  
Dna Polymerase ◽  
Subcellular Fraction ◽  
Polymerase Activity ◽  
Particulate Material ◽  
Newborn Rat ◽  
Newborn Brain ◽  
Rat Brain And Liver ◽  
The Brain

DNA polymerase activities were determined in the cytoplasmic soluble, the nuclear soluble, and the nuclear particulate fractions of newborn rat brain and liver. The results indicate that a majority of the brain nuclear enzyme may be bound to particulate material while a majority of the liver nuclear enzyme may be free or only loosely bound. Although the subcellular distributions of DNA polymerase activity are widely different in newborn brain and liver, the enzyme activity in any given subcellular fraction is higher in liver than in brain.

Formation of yeast mitochondria. II. Effects of antibiotics on enzyme activity during derepression

Biochemistry ◽  
1968 ◽  
Vol 7 (12) ◽  
pp. 4445-4454 ◽  
Author(s):  
Carl P. Henson ◽  
Philip. Perlman ◽  
Celia N. Weber ◽  
Henry R. Mahler
Keyword(s):  
Enzyme Activity ◽  
Yeast Mitochondria

scholarly journals Aphidicolin, an inhibitor of DNA replication, blocks the TPA-induced differentiation of a human megakaryoblastic cell line, MEG-O1

Blood ◽  
1991 ◽  
Vol 78 (12) ◽  
pp. 3168-3177 ◽  
Author(s):  
T Murate ◽  
T Hotta ◽  
K Tsushita ◽  
M Suzuki ◽  
T Yoshida ◽  
...  
Keyword(s):  
Dna Replication ◽  
Cell Growth ◽  
Cell Line ◽  
Cell Lines ◽  
Dna Polymerase ◽  
Phenotypic Expression ◽  
Leukemia Cell ◽  
Specific Inhibitor ◽  
Dna Polymerase Alpha ◽  
Multinuclear Cell

Abstract The commitment process of a human megakaryoblastic cell line (MEG-O1) induced with phorbol ester, TPA, was investigated with special reference to glycoprotein (GP) IIb/IIIa expression, multinuclear formation, and DNA replication. TPA (10(-7) mol/L) completely inhibited cellular division in MEG-O1, but did not suppress de novo DNA synthesis. Two days' culture with 10(-7) mol/L TPA was sufficient for MEG-O1 cells to initiate an irreversible commitment process. These cells could not resume cell growth and expressed GP IIb/IIIa antigen; some of them showed multinuclear form and DNA polyploidy even after removal of TPA from the culture medium. DNA histogram analysis showed that, upon treatment with TPA, the percentage of cells whose DNA ploidy was more than 8N was 5 to 10 times higher than that of control cells. Precise analysis using cell size fractionation by centrifugal elutriation method showed that there was strong correlation between the percentage of multinuclear cells and DNA polyploidy in TPA-treated cells. The percentage and staining intensity of GP IIb/IIIa and other megakaryocytic phenotypes such as von Willebrand factor and PAS staining were highest in large multinuclear cell populations, suggesting that these cells are the most differentiated population in this system. In TPA-treated cells, the activity of DNA polymerase alpha, a marker for cell growth, remained at the same level as in control cells. Aphidicolin, a specific inhibitor of DNA polymerase alpha, completely inhibited the differentiation induction of MEG-O1 cells with TPA measured by either GP IIb/IIIa expression or multinuclear cell formation. Therefore, DNA replication appears to be involved in the process of phenotypic expression as well as endomitosis in megakaryocyte differentiation of MEG-O1 cells. Aphidicolin was also effective in inhibiting megakaryocytic differentiation of other leukemia cell lines such as human erythroleukemia (HEL) and K562 cell lines induced with TPA, suggesting the close interplay of DNA replication and phenotypic expression in megakaryopoiesis.

Interferon-independent activation of (2′-5′) oligoadenylate synthetase in Friend erythroleukemia cell variants exposed to HMBA

1996 ◽  
Vol 109 (6) ◽  
pp. 1517-1526
Author(s):  
S. Salzberg ◽  
A. Heller ◽  
J.P. Zou ◽  
F.R. Collart ◽  
E. Huberman
Keyword(s):  
Enzyme Activity ◽  
Cell Growth ◽  
Oligoadenylate Synthetase ◽  
Down Regulation ◽  
Erythroleukemia Cells ◽  
Regulation Of Activity ◽  
Erythroleukemia Cell ◽  
Cell Variant ◽  
High Treatment ◽  
Induced Proteins

To provide evidence for the implication of interferon (IFN)-induced proteins in the regulation of cell growth during differentiation, the activation of (2′-5′) oligoadenylate synthetase (2-5A synthetase) as well as of PKR, two IFN-induced proteins, during differentiation of Friend erythroleukemia cells, was studied. Two cell variants were used. The first (FL) was completely susceptible to hexamethylene bis-acetamide (HMBA)-treatment and responded in both growth-retardation and hemoglobin synthesis. The second (R1) failed to synthesize hemoglobin in response to HMBA although cell growth was still inhibited. In both cell variants, 2–5A synthetase enzyme activity was induced in a similar fashion, reaching a peak at 26 hours after treatment with HMBA. However, the down regulation of activity thereafter was not identical in both cases. In R1 cells, the reduction was much slower compared to FL cells. A similar pattern was observed with the appearance of the 43 kDa isoform of 2–5A synthetase in immunoblots. An analysis of 2–5A synthetase gene expression revealed the presence of 1.7 kb transcripts which peaked at 16 hours after HMBA-treatment in both cell variants. Again, the down-regulation in expression was slower in R1 than in FL cells. Addition of anti-murin alpha/beta-IFN antibodies did not reduce the level of either 2–5A synthetase expression or enzyme activity in either cell variant. Interestingly, the presence of antibodies also did not affect the pattern of pRb phosphorylation in the cell variants exposed to HMBA. In both cell variants, an increase in the amount of the phosphorylated form (ppRb) was observed in immunoblots after 4 hours. This form was gradually transformed to the underphosphorylated molecule (pRb) with time in culture, even in the presence of antibodies. This further substantiates the notion that IFN-induced regulation of pRb phosphorylation is mediated by IFN-induced proteins. The basal level of either expression or ezymatic activity of PKR detected in untreated FL or R1 cells, was relatively high. Treatment with HMBA did not result in further induction of PKR in either cell variant.

Acute Myeloid Leukemia (AML) Cells Alter the Bone Marrow Microenvironment By Inducing Osteogenic and Suppressing Adipogenic Differentiation of MSCs through BMP-RUNX2-CTGF Mediated Mechanisms

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2403-2403
Author(s):  
Venkata Lokesh Battula ◽  
Phuong M Le ◽  
Jeffrey Sun ◽  
Hong Mu ◽  
Teresa Mc.Queen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Alkaline Phosphatase ◽  
Enzyme Activity ◽  
Cell Growth ◽  
Osteogenic Differentiation ◽  
Research Funding ◽  
Adipogenic Differentiation ◽  
Time Dependent ◽  
Dependent Manner ◽  
Osteoprogenitor Cells

Abstract Osteoprogenitor cells are a critical component of the bone marrow microenvironment (BME), support AML cell growth (Raaijmakers et al., Nature, 2010); however the mechanism has not been fully elucidated. Here, we hypothesize that AML cells induce osteogenic differentiation in mesenchymal stem/stromal cells to gain growth advantage. We have isolated age matched (between 40-70 years) bone marrow derived MSCs from AML patients (AML-MSC, n=20) and normal (N-MSC, n=10) individuals and analyzed their cell growth, cell surface phenotype and multi-lineage differentiation. AML-MSCs are phenotypically different with larger cell surface area and lower cell growth compared to N-MSCs. The average doubling time of AML-MSCs is 52±8hrs compared to 34±6hours for N-MSCs (p<0.01). Phenotypic and enzyme activity studies revealed that tissue non-specific alkaline phosphatase (TNAP), a protein that is highly expressed in naïve MSCs and osteoprogenitor cells and osteoblasts, is expressed 10-14 fold higher in AML-compared to N-MSCs (p<0.01). As TNAP is associated with osteogenic lineage, we next assessed the expression of osteogenic markers including RUNX-2, osteopontin, TNAP and osterix. Interestingly, these genes were up-regulated by 5-10-fold in AML-MSCs compared to N-MSCs. To validate these observations, N-MSCs were cultured with OCI-AML3 cell derived conditioned medium (CM) for 3-5 days and then induced to osteogenic or adipogenic differentiation. As expected, alkaline phosphatase enzyme activity and Alizarine Red S staining was twice as high in MSCs cultured with AML-CM compared to MSCs cultured in regular cell culture medium. In addition, we found expression of osteogenic genes including RUNX-2, osteopontin, TNAP and osterix 3-4 fold upregulated in MSCs cultured with AML-CM compared to control MSCs. These data indicate that AML cells prime MSCs to undergo osteogenic differentiation. Adipocyte differentiation was assessed by Oil-Red O staining for lipid droplets and revealed a >95% reduction (p<0.001) in the number of mature adipocytes in AML-MSCs compared to N-MSCs. Gene expression analysis by qRT-PCR revealed that adipogenic markers including aP2, lipoprotein lipase and PPARγ were down-regulated 10-20 fold in AML-MSCs compared to N-MSCs in a time-dependent manner suggesting that AML-MSCs lack the ability to differentiate into adipocytes. To evaluate these findings in-vivo, we developed a Human Bone Marrow Implant (HBMI) mouse model by subcutaneous implantation of human femur derived bone pieces into NSG mice. In this model, mice bearing HBMI with leukemia expressed higher osteogenic related proteins compared to mice without leukemia. Importantly, multispectral image analysis revealed 5-10 fold higher osteogenic activity in AML patient BM-biopsies compared to normal bones. Bone morphogenic proteins (BMP) are the most essential factors during osteogenic differentiation and new bone formation in humans. We therefore treated MSCs with AML cell-derived conditioned medium, and observed a 6-8 fold increase in pSmad1/5 levels in N-MSCs in a time dependent manner. In addition, AML induced pSmad1/5 up-regulation was inhibited when MSCs were treated with BMP-type1 receptor specific inhibitor LDN-212854, in a dose dependent manner. In addition, inhibition of BMP signaling also inhibited AML-induced osteogenic differentiation in MSCs. Mechanistic studies revealed that bone morphogenetic proteins (BMPs) derived from AML cells induce connective tissue growth factor (CTGF) expression in MSCs. When transplanted in Col1a2-CTGF transgenic mice, AML cells engrafted faster compared to wild type mice suggesting that CTGF enhances AML cell growth. Our data suggest that AML cells induce osteogenic differentiation in MSCs and receive growth advantage through BMP-CTGF signaling. Inhibition of these pathways may reduce leukemia propagation and improve AML patient survival. Disclosures Konopleva: Novartis: Research Funding; AbbVie: Research Funding; Stemline: Research Funding; Calithera: Research Funding; Threshold: Research Funding.

Biochemical, cellular and molecular identification of DNA polymerase α in yeast mitochondria

Biochimie ◽  
2013 ◽  
Vol 95 (4) ◽  
pp. 759-771 ◽  
Author(s):  
Jean-Paul Lasserre ◽  
Jacqueline Plissonneau ◽  
Christophe Velours ◽  
Marc Bonneu ◽  
Simon Litvak ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Molecular Identification ◽  
Yeast Mitochondria ◽  
Dna Polymerase Α

Regulation of eukaryotic DNA replication at the initiation step

2003 ◽  
Vol 31 (1) ◽  
pp. 266-269 ◽  
Author(s):  
S. Pollok ◽  
J. Stoepel ◽  
C. Bauerschmidt ◽  
E. Kremmer ◽  
H.-P. Nasheuer
Keyword(s):  
Dna Replication ◽  
Cell Growth ◽  
Dna Polymerase ◽  
Molecular Control ◽  
Dna Polymerase Α ◽  
Late Protein ◽  
Two Factors ◽  
Eukaryotic Dna ◽  
A Cell ◽  
The Stability

The studies of cell growth and division have remained at the centre of biomedical research for more than 100 years. The combination of genetic, biochemical, molecular and cell biological techniques recently yielded a burst in what is known of the molecular control of cell growth processes. The initiation of DNA replication is crucial for the stability of the genetic information of a cell. Two factors, Cdc45p (cell division cycle 45p) and DNA polymerase α-primase, are necessary in this process. Depending on growth signals, Cdc45p is expressed as a late protein. New phosphorylation-specific antibodies specifically recognize the phosphorylated subunit, p68, of the four subunit DNA polymerase α-primase and show that the phosphorylated polypeptide is exclusively nuclear.

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