scholarly journals Accumulation of p53 in a mutant cell line defective in the ubiquitin pathway.

1994 ◽  
Vol 14 (3) ◽  
pp. 1997-2003 ◽  
Author(s):  
D R Chowdary ◽  
J J Dermody ◽  
K K Jha ◽  
H L Ozer
Keyword(s):  
Cell Line ◽  
Mutant Cell ◽  
P53 Gene ◽  
Mouse Cell ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Wild Type ◽  
Proteolytic Pathway ◽  
Ubiquitination Pathway ◽  
Wild Type P53

The wild-type p53 gene product plays an important role in the control of cell proliferation, differentiation, and survival. Altered function is frequently associated with changes in p53 stability. We have studied the role of the ubiquitination pathway in the degradation of p53, utilizing a temperature-sensitive mutant, ts20, derived from the mouse cell line BALB/c 3T3. We found that wild-type p53 accumulates markedly because of decreased breakdown when cells are shifted to the restrictive temperature. Introduction of sequences encoding the human ubiquitin-activating enzyme E1 corrects the temperature sensitivity defect in ts20 and prevents accumulation of p53. The data therefore strongly indicate that wild-type p53 is degraded intracellularly by the ubiquitin-mediated proteolytic pathway.

scholarly journals Accumulation of p53 in a mutant cell line defective in the ubiquitin pathway

1994 ◽  
Vol 14 (3) ◽  
pp. 1997-2003
Author(s):  
D R Chowdary ◽  
J J Dermody ◽  
K K Jha ◽  
H L Ozer
Keyword(s):  
Cell Line ◽  
Mutant Cell ◽  
P53 Gene ◽  
Mouse Cell ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Wild Type ◽  
Proteolytic Pathway ◽  
Ubiquitination Pathway ◽  
Wild Type P53

The wild-type p53 gene product plays an important role in the control of cell proliferation, differentiation, and survival. Altered function is frequently associated with changes in p53 stability. We have studied the role of the ubiquitination pathway in the degradation of p53, utilizing a temperature-sensitive mutant, ts20, derived from the mouse cell line BALB/c 3T3. We found that wild-type p53 accumulates markedly because of decreased breakdown when cells are shifted to the restrictive temperature. Introduction of sequences encoding the human ubiquitin-activating enzyme E1 corrects the temperature sensitivity defect in ts20 and prevents accumulation of p53. The data therefore strongly indicate that wild-type p53 is degraded intracellularly by the ubiquitin-mediated proteolytic pathway.

Correction of the defect in initiation of DNA replication in a temperature-sensitive mutant hamster cell line by in vitro addition of extracts from normal cells

1985 ◽  
Vol 5 (4) ◽  
pp. 902-905
Author(s):  
M Narkhammar ◽  
R Hand
Keyword(s):  
Cell Line ◽  
Nuclear Extract ◽  
Cell Extract ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Permissive Temperature ◽  
Wild Type ◽  
Whole Cell ◽  
Mutant Cells

ts BN-2 is a temperature-sensitive hamster cell line that is defective in DNA synthesis at the restrictive temperature. The mutant expresses its defect during in vitro replication in whole-cell lysates. Addition of a high-salt-concentration extract from wild-type BHK-21, revertant RBN-2, or CHO cells to mutant cells lysed with 0.01% Brij 58 increased the activity in the mutant three- to fourfold, so that it reached 85% of the control value, and restored replicative synthesis. The presence of extract had an insignificant effect on wild-type and revertant replication and on mutant replication at the permissive temperature. Extract prepared from mutant cells was less effective than the wild-type cell extract was. Also, the stimulatory activity was more heat labile in the mutant than in the wild-type extract. Nuclear extract was as active as whole-cell extract.

scholarly journals Restoration of endogenous wild-type p53 activity in a glioblastoma cell line with intrinsic temperature-sensitive p53 induces growth arrest but not apoptosis

2001 ◽  
Vol 94 (1) ◽  
pp. 35-43 ◽  
Author(s):  
Jun Ikeda ◽  
Mitsuhiro Tada ◽  
Nobuaki Ishii ◽  
Hideyuki Saya ◽  
Kazuhiko Tsuchiya ◽  
...  
Keyword(s):  
Cell Line ◽  
Growth Arrest ◽  
Temperature Sensitive ◽  
Glioblastoma Cell Line ◽  
Glioblastoma Cell ◽  
Wild Type ◽  
Wild Type P53

Exogenous Wild-Type p53 Gene Improved Survival of Nude Mice Injected with Murine Erythroleukemia Cell Line Through Amelioration of Hemorheological Changes

2007 ◽  
Vol 14 (2) ◽  
pp. 155-166 ◽  
Author(s):  
Yingyu Zhang ◽  
Weijuan Yao ◽  
Zhu Zeng ◽  
Xianwei Wang ◽  
Dagong Sun ◽  
...  
Keyword(s):  
Cell Line ◽  
Nude Mice ◽  
P53 Gene ◽  
Wild Type ◽  
Erythroleukemia Cell ◽  
Wild Type P53 ◽  
Murine Erythroleukemia ◽  
Murine Erythroleukemia Cell

scholarly journals Correction of the defect in initiation of DNA replication in a temperature-sensitive mutant hamster cell line by in vitro addition of extracts from normal cells.

1985 ◽  
Vol 5 (4) ◽  
pp. 902-905 ◽  
Author(s):  
M Narkhammar ◽  
R Hand
Keyword(s):  
Cell Line ◽  
Nuclear Extract ◽  
Cell Extract ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Permissive Temperature ◽  
Wild Type ◽  
Whole Cell ◽  
Mutant Cells

ts BN-2 is a temperature-sensitive hamster cell line that is defective in DNA synthesis at the restrictive temperature. The mutant expresses its defect during in vitro replication in whole-cell lysates. Addition of a high-salt-concentration extract from wild-type BHK-21, revertant RBN-2, or CHO cells to mutant cells lysed with 0.01% Brij 58 increased the activity in the mutant three- to fourfold, so that it reached 85% of the control value, and restored replicative synthesis. The presence of extract had an insignificant effect on wild-type and revertant replication and on mutant replication at the permissive temperature. Extract prepared from mutant cells was less effective than the wild-type cell extract was. Also, the stimulatory activity was more heat labile in the mutant than in the wild-type extract. Nuclear extract was as active as whole-cell extract.

scholarly journals Wild-type p53 gene expression induces granulocytic differentiation of HL-60 cells

Blood ◽  
1994 ◽  
Vol 83 (8) ◽  
pp. 2230-2237 ◽  
Author(s):  
S Soddu ◽  
G Blandino ◽  
G Citro ◽  
R Scardigli ◽  
G Piaggio ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cellular Response ◽  
P53 Protein ◽  
Cell Cycle Control ◽  
P53 Gene ◽  
Malignant Cell ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Granulocytic Differentiation ◽  
Wild Type P53

Abstract Overexpression of wild-type p53 gene in malignant cell lines has been shown to inhibit cell proliferation in a number of cases. However, endogenous p53 protein seems to play little role in normal cell-cycle control as suggested by the normal development of p53 null mice, and by the low p53 protein levels expressed in most cell types. Recently, increased expression of endogenous p53 protein has been observed during the cellular response to DNA damage, as well as during differentiation of human hematopoietic cells. To study the role of the p53 gene in hematopoietic differentiation, we introduced the wild-type p53 gene or the temperature-sensitive p53(Val135) mutant into p53-deficient HL-60 promyelocytic leukemia cells. Morphological analysis, flow-cytometric determination of granulocytic or monocytic surface markers, and ability to reduce nitroblue tetrazolium (NBT) demonstrated that expression of exogenous wild-type p53 gene in HL-60 cells induces differentiation through the granulocytic pathway. Proliferation and cell-cycle analysis performed early after expression of wild-type p53 showed that induction of differentiation is not coupled with growth arrest, which suggests that p53 is involved in differentiation independently of its activity on the cell cycle.

scholarly journals Analysis of p53 mutants for transcriptional activity.

1991 ◽  
Vol 11 (12) ◽  
pp. 6067-6074 ◽  
Author(s):  
L Raycroft ◽  
J R Schmidt ◽  
K Yoas ◽  
M M Hao ◽  
G Lozano
Keyword(s):  
Transcription Factor ◽  
Fusion Protein ◽  
Fusion Proteins ◽  
P53 Protein ◽  
P53 Gene ◽  
Temperature Sensitive ◽  
Mutant P53 ◽  
Permissive Temperature ◽  
Wild Type ◽  
Wild Type P53

The wild-type p53 protein functions to suppress transformation, but numerous mutant p53 proteins are transformation competent. To examine the role of p53 as a transcription factor, we made fusion proteins containing human or mouse p53 sequences fused to the DNA binding domain of a known transcription factor, GAL4. Human and mouse wild-type p53/GAL4 specifically transactivated expression of a chloramphenicol acetyltransferase reporter in HeLa, CHO, and NIH 3T3 cells. Several mutant p53 proteins, including a mouse p53 mutant which is temperature sensitive for suppression, were also analyzed. A p53/GAL4 fusion protein with this mutation was also transcriptionally active only at the permissive temperature. Another mutant p53/GAL4 fusion protein analyzed mimics the mutation inherited in Li-Fraumeni patients. This fusion protein was as active as wild-type p53/GAL4 in our assay. Two human p53 mutants that arose from alterations of the p53 gene in colorectal carcinomas were 30- to 40-fold less effective at activating transcription than wild-type p53/GAL4 fusion proteins. Thus, functional wild-type p53/GAL4 fusion proteins activate transcription, while several transformation competent mutants do so poorly or not at all. Only one mutant p53/GAL4 fusion protein remained transcriptionally active.

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