PTEN, a tumor suppressor commonly targeted in human cancer, possesses phosphatase activities toward both protein and lipid substrates. While PTEN suppresses gliomas through cell cycle inhibition which requires its lipid phosphatase activity, PTEN's effects on other tumor types and the role of its protein phosphatase activity are controversial or unknown. Here we show that exogenous wild-type PTEN arrests some, but not all human breast cancer cell lines in G1, in a manner independent of endogenous PTEN. Unexpectedly, the G129E mutant of PTEN selectively deficient in the lipid phosphatase activity still blocked the cell cycle of MCF-7 cells, while the G129R and H123Y mutants lacking both phosphatase activities were ineffective. These results suggest that PTEN's protein phosphatase activity likely contributes to its tumor suppressor function in subsets of tumors and that elucidation of downstream targets which dictate cellular responses to PTEN may have important implications for future cancer treatment strategies. Copyright 2000 Academic Press.
- MeSH
- buněčný cyklus MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- fosfatasy fyziologie genetika MeSH
- fosfohydroláza PTEN MeSH
- lidé MeSH
- mutace MeSH
- nádorové buňky kultivované MeSH
- nádorové supresorové proteiny * MeSH
- proteinfosfatasy * metabolismus MeSH
- tumor supresorové geny * MeSH
- western blotting MeSH
- Check Tag
- lidé MeSH
p27Kip1 is a regulator of the mammalian cell cycle and a putative tumor suppressor. Distinct altered patterns of p27Kip1 protein expression are found in a variety of human carcinomas, and p27Kip1 expression levels usually correlate directly with disease-free survival. The mechanism(s) by which p27Kip1 expression is reduced or lost during tumorigenesis remains unclear. Specific alterations of the p27Kip1 gene, including mutations and homozygous deletions, are exceedingly rare in human cancers. We have used methylation-specific PCR and bisulfite genomic sequencing to examine the methylation status of p27Kip1 in 61 primary and metastatic tumors and 35 cell lines from patients with malignant melanoma. Dense methylation of a CpG island in the promoter region of p27Kip1 was detected in four of 45 metastatic tumors (9%) and three of the cell lines (9%), including two cell lines established from two different metastases from the same patient. Examination of a naturally occurring, allele-specific sequence variant demonstrated that p27Kip1 methylation is associated with transcriptional silencing in situ. Cell lines with p27Kip1 methylation showed retention of the wild-type allele and detectable p27Kip1 protein whose abundance was reduced compared with normal melanocytes. Collectively, our data suggest that DNA methylation may be a cause of monoallelic p27Kip1 silencing in malignant melanoma, which would support a role for p27Kip1 haploinsufficiency in human cancer.
- MeSH
- aktivace transkripce MeSH
- CpG ostrůvky fyziologie MeSH
- inhibitor p27 cyklin-dependentní kinasy MeSH
- lidé MeSH
- melanocyty metabolismus MeSH
- melanom * genetika metabolismus MeSH
- messenger RNA biosyntéza genetika MeSH
- metylace DNA * MeSH
- mutace MeSH
- nádorové supresorové proteiny * MeSH
- polymerázová řetězová reakce MeSH
- promotorové oblasti (genetika) genetika MeSH
- proteiny asociované s mikrotubuly * genetika metabolismus MeSH
- proteiny buněčného cyklu * MeSH
- regulace genové exprese u nádorů genetika MeSH
- substituce aminokyselin MeSH
- umlčování genů MeSH
- Check Tag
- lidé MeSH
- MeSH
- adenosintrifosfatasy MeSH
- antigeny MeSH
- cytotoxické T-lymfocyty MeSH
- melanom imunologie MeSH
- Publikační typ
- přehledy MeSH
- MeSH
- antigeny CD80 genetika MeSH
- buněčná adheze MeSH
- experimentální sarkom imunologie patofyziologie MeSH
- genetická terapie MeSH
- modely nemocí na zvířatech MeSH
- myši MeSH
- protinádorové vakcíny genetika imunologie MeSH
- T-lymfocyty fyziologie MeSH
- techniky in vitro MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
Two genes, the gene coding for IL-2 and the gene encoding the CD80 molecule, were inserted into murine sarcoma MC12 cells. Tumorigenicity of a variety of cell clones with different expression of the inserted genes was assessed. Most of the genetically manipulated MC12 cell clones were less tumorigenic than the parental MC12 cell population. Tumorigenicity of the clones declined with increasing production of IL-2 as well as with the increasing expression of the CD80 molecule. When the tumorigenicity of the clones carrying an inserted IL-2 gene was compared with that of the clones carrying an inserted CD80 gene, it was found that the insertion of the IL-2 gene suppresses tumorigenicity more efficiently than insertion of the CD80 gene. Admixture of the IL-2-producing MC12 clones to the tumorigenic CD80(+) MC12 cell doses could completely inhibit the tumorigenicity of the CD80(+) cells. Insertion of the CD80 gene into sarcoma cells substantially enhanced the adhesive interaction between the MC12 sarcoma and syngeneic T lymphocytes.
- MeSH
- antigeny CD80 MeSH
- antitumorózní látky MeSH
- interleukin-2 MeSH
- myši MeSH
- protinádorové vakcíny MeSH
- sarkom genetika MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- práce podpořená grantem MeSH
The p16Ink4/CDKN2, D-type cyclins, their partners Cdk4/Cdk6, and pRb constitute a G1 regulatory pathway commonly targeted in tumorigenesis. Genetic, immunochemical, and functional cell cycle analyses showed abnormalities of this pathway in each of 22 human melanoma cell lines examined. Normal melanocytes and all melanoma lines expressed Cdk4, Cdk6, and cyclins D1 and D3. The tumor suppressors p16Ink4/CDKN2 and pRb were lost in 17 and 4 cases, respectively, due to various genetic mechanisms, including transcriptional block of p16 and nonsense mutations of RB1. Ectopic expression of p16 prevented S-phase entry of Rb+/p16- but not Rb-deficient melanoma lines. The SK29-MEL-1 cell line harboring an R24C mutation in Cdk4 expressed wild-type pRb and overabundant p16, the latter preventing endogenous Cdk6 but not Cdk4 from associating with cyclin D1. Microinjection of cyclin D1-neutralizing antibody arrested the SK29-MEL-1 cells in G1, whereas pl6 did not, indicating that the cyclin D1/Cdk4-R24C complex is required for G1 progression, and the resistance of the complex to p16 in vivo. These data strongly support the candidacy of Cdk4 as a novel proto-oncogene, provide further evidence for the p16-cyclin D/Cdk-pRb pathway as a functional unit, and suggest that deregulation of this checkpoint may represent a common step in the multistep progression of sporadic malignant melanomas.
- MeSH
- cyklin D MeSH
- cyklin-dependentní kinasa 4 MeSH
- cyklin-dependentní kinasy genetika fyziologie MeSH
- cykliny genetika fyziologie MeSH
- delece genu MeSH
- G1 fáze fyziologie MeSH
- geny retinoblastomu MeSH
- inhibitor p16 cyklin-dependentní kinasy MeSH
- lidé MeSH
- melanom etiologie genetika patofyziologie MeSH
- molekulární sekvence - údaje MeSH
- nádorová transformace buněk genetika MeSH
- nádorové buňky kultivované MeSH
- polymerázová řetězová reakce MeSH
- progrese nemoci MeSH
- protoonkogenní proteiny * MeSH
- protoonkogeny MeSH
- retinoblastomový protein fyziologie MeSH
- sekvence nukleotidů MeSH
- signální transdukce fyziologie MeSH
- transportní proteiny genetika fyziologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- buněčná imunita MeSH
- buňky NK cytologie MeSH
- genetická terapie metody MeSH
- interleukin-2 genetika terapeutické užití MeSH
- myši MeSH
- plazmocytom imunologie terapie MeSH
- rekombinantní proteiny terapeutické užití MeSH
- T-lymfocyty cytologie MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
