Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.22 (cdc2)
 8,319 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In vitro cell transformation is a valuable approach for studying the mechanisms of multistep carcinogenesis of human cells. Since immortalization is an essential step for in vitro neoplastic transformation of human cells, this study addresses the question of whether mutant p53 contributes to the immortalization process of human cells. The mutant p53 gene (mp53: codon273Arg-His) was introduced into normal human fibroblasts (OUMS-24 line) and a G418-resistant clone, OUMS-24/P6 line, was obtained. This clone showed an extended life span and chromosome abnormalities, but senesced at the 79th population doubling level (PDL). When these cells were subjected to intermittent X-ray treatment, they became an immortalized cell line (OUMS-24/P6X). Although these immortalized cells showed chromosome abnormalities, they were not tumorigenic. On the other hand, normal OUMS-24 cells into which mp53 had not been introduced were not immortalized by the same X-ray treatment. These results indicate that introduction and expression of mp53 alone were not sufficient for immortalization of human cells, and that mutations of the remaining wild-type p53 or other genes may have been necessary for immortalization. In fact, no expression of the wild-type p53 was detected in the immortalized cells by RT-PCR. Expression of p21, which is located downstream of p53, was remarkably reduced in the immortalized cells, resulting in an increase in cdk2 and cdc2 kinase activity. These findings indicate that the p53-p21 cascade may play some role in the immortalization of human cells. On the other hand, there was no significant difference in expression of proteins such as Rb, p16, cdk4, cdk6, cyclin A and cyclin D1 between the normal and immortalized human fibroblasts.
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PMID:Transformation of normal human fibroblasts into immortalized cells with the mutant p53 gene and X-rays. 898 2

P130 shares structural and functional homology with pRb and p107. One property common to p107 and p130, but not to pRb, is the ability to stably interact with cyclin A/cdk2 and cyclin E/cdk2 complexes in vitro and in vivo. Using GST-p130 fusion proteins representing various regions of p130, baculovirus-produced cyclin A/cdk2 and cyclin E/cdk2 complexes were found to interact with residues within a part of p130 known as the spacer region. Cyclin E was able to bind the p130 spacer region in the presence or absence of cdk2 whereas cyclin A binding was dependent upon the presence of cdk2. The smallest p130 fusion protein sufficient to interact with cyclin A/cdk2 or cyclin E/cdk2 complexes contained p130 amino acids 652-698 and deletion of p130 amino acids 680-682 abolished binding to both of the cyclin/cdk2 complexes. When overexpressed in C33A cells, a p130 mutant containing a deletion of amino acids 620-697 was unable to form complexes with either cyclin A or cyclin E. This p130 mutant was at least as active as wild type p130 in suppressing the growth of G418 resistant colonies when overexpressed in C33A or SAOS-2 cells.
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PMID:Identification of a p130 domain mediating interactions with cyclin A/cdk 2 and cyclin E/cdk 2 complexes. 918 54

The study of in vitro cell transformation is valuable for understanding the multistep carcinogenesis of human cells. The difficulty in inducing neoplastic transformation of human cells by treatment with chemical or physical agents alone is due to the difficulty in immortalizing normal human cells. Thus, the immortalization step is critical for in vitro neoplastic transformation of human cells. We transfected a mutant p53 gene (mp53: codon 273Arg-His) into normal human fibroblasts and obtained two G418-resistant mp53-containing clones. These clones showed an extended life span but ultimately senesced. However, when they were treated with either 4-nitroquinoline 1-oxide or X-rays, they were immortalized. The immortalized cells showed both numerical and structural chromosome abnormalities, but they were not tumorigenic. The expression of mutant but not wild type p53 was detected in the immortalized cells by RT-PCR. Expression of p21, which is located downstream of p53, was remarkably reduced in the immortalized cells, resulting in increased cdk2 and cdc2 kinase activity. However, there was no significant difference between the normal and immortalized human cells in expression of another tumor suppressor gene, p16. These findings indicate that the p53-p21 cascade may play an important role in the immortalization of human cells.
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PMID:Immortalization of mutant p53-transfected human fibroblasts by treatment with either 4-nitroquinoline 1-oxide or X-rays. 933 45

To duplicating the regulatory subunit p35Nck5a gene of mouse neuronal cdc2-like kinase in embryonic stem (ES) cells, about 12.2 kb of pGDTV vector for p35Nck5a gene duplication was constructed. The linearized pGDTV vectors were transfected into ES cells by electroporation. 245 drug-resistant cell clones were obtained in both G418 and GANC medium and the frequency of surviving cells was 6.22 x 10(-5). The ES cell clones were identified to have duplicated the p35Nck5a gene by use of both PCR and genomic Southern blotting, and the frequency of homologous recombination is 5.08 x 10(-7). The use of negative selection gene (HSV-tk) results in 7-fold increase at selection efficiency. This study lays the foundations of preparing mouse models of Alzheimer's disease.
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PMID:[Construction of gene targeting vector for duplicating p35Nck5a gene and its application in the gene targeting of ES cells]. 1105 17

To study the effect on regulation of cell cycle of osteosarcoma cell line MG63 tranceduced with exogenous p16ink4a and hRb1 genes, pIRES-p16ink4a-hRb1, pIRES-p16ink4a and pIRES-hRb1 plasmids were constructed by gene recombination technology. The recombinant plasmid was transferred into osteosarcoma cell line MG63 by metafectene, and the resistant clones were selected by G418 selective medium. mRNA and protein expression of osteosarcoma cell line were assayed by RT-PCR and Western-Blot respectively. Cell cycle and apoptosis were analyzed by subG1 flow cytometric. Cell proliferation was tested by MTT. In the genome of these transfected target cells, the expression of p16ink4a and hRb1 mRNA and protein were detected respectively in vitro. It was demonstrated with subG1 flow cytometric analysis and MTT method that p16ink4a and hRb1 genes cooperation more significantly inhibited cell growth and induced a more marked G1 arrest and apoptosis than p16ink4a/hRb1 alone (P < 0.01). Coexpression of exogenous p16ink4a with hRb1 broke the regulatory feedback loop of p16ink4a-cyclinD1 /CDK-hRb1 and played a more significant role in inhibiting cell growth as well as inducing cell apoptosis than p16ink4a or hRb1 did alone in vitro.
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PMID:Effect of exogenous p16ink4a and hRb1 genes on cell cycle regulation of osteosarcoma cell. 1669 24