The RAS oncogene induces genomic instability in thyroid PCCL3 cells via the MAPK pathway.
Activating mutations of RAS are thought to be early events in the evolution of thyroid follicular neoplasms. We used a doxycycline-inducible expression system to explore the acute effects of H-RAS12 on genomic stability in thyroid PCCL3 cells. At 2-3 days (first or second cell cycle) there was a significant increase in the frequency of micronucleation. Treatment of cells with YVAD-CHO inhibited RAS-induced apoptosis, but had no effect on micronucleation. The effects of H-RAS(V12) were mediated by activation of MAPK, as treatment with PD98059 at concentrations verified to selectively inhibit MEK1 reduced the frequency of prevalence of cells with micronuclei. In addition, doxycycline-inducible expression of a constitutively active MEK1, but not of a mutant RAC1, mimicked the effects of H-RAS(V12). The effects of H-RAS(V12) on genome destabilization were apparent even though the sequence of p53 in PCCL3 cells was confirmed to be wild-type. Acute activation of H-RAS(V12) evoked a proportional increase in both CREST negative and CREST positive micronuclei, indicating that both clastogenic and aneugenic effects were involved. H-RAS(V12) and activated MEK1 also induced centrosome amplification, and chromosome misalignment. Evidence that acute expression of constitutively activated RAS destabilizes the genome of PCCL3 cells is consistent with a mode of tumor initiation in which this oncogene promotes phenotypic progression by predisposing to large scale genomic abnormalities.
Saavedra
H I
HI
Division of Endocrinology and Metabolism, University of Cincinnati College of Medicine, Ohio 45267, USA.
Knauf
J A
JA
Shirokawa
J M
JM
Wang
J
J
Ouyang
B
B
Elisei
R
R
Stambrook
P J
PJ
Fagin
J A
JA
D000818:Animals; D017209:Apoptosis / Q000528:radiation effects; D002467:Cell Nucleus / Q000187:drug effects / Q000235:genetics; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D004318:Doxycycline / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D005720:Gamma Rays; D015972:Gene Expression Regulation, Neoplastic; D048369:MAP Kinase Kinase 1; D020935:MAP Kinase Signaling System; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors / Q000235:genetics; D009154:Mutation; D010749:Phosphoprotein Phosphatases / Q000235:genetics; D054648:Protein Phosphatase 2; D000071636:Protein Phosphatase 2C; D017346:Protein Serine-Threonine Kinases / Q000235:genetics; D051381:Rats; D029701:Saccharomyces cerevisiae Proteins; D013964:Thyroid Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000378:metabolism; D014407:Tumor Cells, Cultured; D020830:rac1 GTP-Binding Protein / Q000235:genetics; D018631:ras Proteins / Q000235:genetics*
D002868:Chromosomal Proteins, Non-Histone; D004791:Enzyme Inhibitors; D005419:Flavonoids; D029701:Saccharomyces cerevisiae Proteins; C068688:centromere protein E; D017346:Protein Serine-Threonine Kinases; D048369:MAP Kinase Kinase 1; D020929:Mitogen-Activated Protein Kinase Kinases; C087358:PTC1 protein, S cerevisiae; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D000071636:Protein Phosphatase 2C; D020830:rac1 GTP-Binding Protein; D018631:ras Proteins; D004318:Doxycycline; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one