RAS oncogene-mediated deregulation of the transcriptome: From molecular signature to function (original) (raw)
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Biomarkers downstream of RAS: a search for robust transcriptional targets
2010
The small GTP-binding proteins HRAS, KRAS and NRAS belong to a family of oncoproteins associated with many types of human cancer. Signal transduction processes initiated at receptor tyrosine kinases converge on RAS proteins which serve as molecular switches linking upstream signals with the transcriptional machinery. RAS proteins interact with a number of effector proteins that in turn activate the Raf/MEK/ERK pathway, the PI3K/PKB/Akt pathway, the RalGDS/Ral pathway and other downstream pathways. Mutations in RAS lock the protein in its active form. Chronic activation of the KRAS isoform is the basis for resistance toward antibody therapies targeting receptor tyrosine kinases, as an upstream stimulus through growth factor receptor-mediated activation is no longer required. However, the complexity of the RAS signaling system necessitates the search for additional activating mechanisms as well as biomarkers associated with pathway activation. During recent years, several RAS pathway-related gene signatures were identified, mostly by microarray-based gene expression profiling of normal versus RAS-transformed cells. The signatures can serve as a source of common biomarkers indicating functionally relevant downstream effects of the RAS signaling system. In searching for new markers, we compared the gene expression signatures compiled in 24 independent studies. We analyzed differentially regulated genes recovered in microarray studies on human specimens to discriminate paired normal and tumor tissues. Although the overlap between individual studies was low, this meta-analysis revealed Kruppel-like factor 5 (KLF5), the CD44 antigen and members of the epidermal growth factor (EGR)-family as common downstream effectors of RAS.
Positive autoregulation of ras genes expression in fibroblasts
FEBS Letters, 1997
We have studied the effect of ectopie overexpression of a ras gene on the expression of the other two members of the ras gene family. We obtained NIH3T3 cell lines stably transfected with inducible H-ras and N-ras oncogenes. The expression of these genes is driven by a glucocorticoid-responsive promoter and the addition of dexamethasone resulted in a dramatic induction (10-20-fold) of H-or N-ras mRNA, peaking 4 h after hormone addition. The induction of the expression of ras oncogenes resulted in a transformed phenotype. In quiescent NIH3T3 cells transfected with inducible H-ras oncogenes, the induction of H-Ras was followed 12 h later by a 3-fold increase in the mRNA expression of endogenous K-ras and N-ras. Similarly, in NIH3T3 transfected with inducible N-ras oncogene, the induction of N-ras was followed by an increase in the expression of endogenous K-and H-ras genes. Interestingly, the effect was not limited to the mutated N-ras, as a similar result was obtained in cells transfected with N-ras proto-oncogene. The induction of ras genes expression was not linked to cell cycle progression as it was reproduced in cells arrested in S-phase by pretreatment with hydroxyurea. These results suggest the presence of a positive cross-regulation in the expression among the members of the Ras family. This effect could play a role in Ras-mediated carcinogenesis.
Functional specific roles of H-ras and N-ras. A proteomic approach using knockout cell lines
ELECTROPHORESIS, 2012
Ras small GTPases function as transducers of extracellular signals regulating cell survival, growth and differentiation. There are three major ras isoforms: H-, N-and K-Ras. To improve the understanding of Hand N-Ras protein signalling networks, we compared total proteome changes in mouse embryonic fibroblasts knock out for H-ras and/or N-ras, using proteomics tools combining 2DE, semi-quantitative image analysis, in-gel trypsin digestion and mass spectrometry. There are four up-regulated proteins due to the loss of expression of H-Ras (including cyclin-dependent kinase inhibitor 2A) and eight downregulated (including stress-70 protein, dihydropyrimidinase-related-protein 3, heat shock cognate 71 kDa protein, tropomyosin beta chain, Rho GDP-dissociation inhibitor 1) and six up-regulated proteins (e.g. leukocyte elastase inhibitor A, L-lactate dehydrogenase B chain, c-Myc-responsive protein Rcl, interleukin-1 receptor antagonist protein) due to the loss of expression of both N-and H-Ras. Most of these proteins are related to Ras signalling in one way or another. Changes in expression of some of these proteins were further confirmed by Western blot. This proteomic comparative analysis from loss of function of Hand N-Ras knockout fibroblasts yields interpretable data to elucidate the differential protein expression, and contributes to evaluate the possibilities for physiological and therapeutic targets.
Ras isoform abundance and signalling in human cancer cell lines
Oncogene, 2008
The ubiquitously expressed major Ras isoforms: H-, Kand N-Ras, are highly conserved, yet exhibit different biological outputs. We have compared the relative efficiencies with which epidermal or hepatocyte growth factor activates Ras isoforms and the requirement for specific isoforms in the activation of downstream pathways. We find that the relative coupling efficiencies to each Ras isoform are conserved between stimuli. Furthermore, in both cases, inhibition of receptor endocytosis led to reduced N-and H-Ras activation, but K-Ras was unaffected. Acute knockdown of each isoform with siRNA allows endogenous Ras isoform function and abundance to be probed. This revealed that there is significant variation in the contribution of individual isoforms to total Ras across a panel of cancer cell lines although typically KXNbH. Intriguingly, cancer cell lines where a significant fraction of endogenous Ras is oncogenically mutated showed attenuated activation of canonical Ras effector pathways. We profiled the contribution of each Ras isoform to the total Ras pool allowing interpretation of the effect of isoform-specific knockdown on signalling outcomes. In contrast to previous studies indicating preferential coupling of isoforms to Raf and PtdIns-3kinase pathways, we find that endogenous Ras isoforms show no specific coupling to these major Ras pathways.
Cancer informatics, 2017
The 3 human RAS genes play pivotal roles regulating proliferation, differentiation, and survival in normal cells and become mutated in 15% to 20% of all human tumors and amplified in many others. In this report, we examined data from The Cancer Genome Atlas to investigate the relationship between RAS gene mutational status and messenger RNA expression. We show that all 3 RAS genes exhibit increased expression when they are mutated in a context-dependent manner. In the case of KRAS, this increase is manifested by a larger proportional increase in KRAS4A than KRAS4B, although both increase significantly. In addition, the mutational status of RAS genes can be associated with expression changes in other RAS genes, with most of these cases showing decreased expression. The mutational status associations with expression are recapitulated in cancer cell lines. Increases in expression are mediated by both copy number variation and contextual differences, including mutational status of epide...
K-ras expression--marker of dysplasia and cancer
Roumanian archives of microbiology and immunology
Molecular and epidemiological data indicated that the presence of HPV virus is not sufficient to induce transformation, suggesting the implication of other several cellular factors. Constitutive activation of the Ras signaling pathway is an important component of malignant progression for a number of different cancers. In this context, the objectives of our study were: the quantitative assessment of the K-ras gene expression changes in the development of the HPV positive cervical cancers. We observed that the K-ras mRNA expression levels did not gradually increase with the severity of injury. The mRNA expression in the ASCUS increased 2.02 times as compared with the control group, while in LSIL group only 1.76 times. However ras expression was increased in the HSIL/cancer group by 2.27 times when was reported to the control group. The presence of low risk HPV infection (IrHPV) does not lead to increased ras expression, remaining at baseline, but K-ras expression was increased in the...
Distinct requirements for Ras oncogenesis in human versus mouse cells
Genes & Development, 2002
The spectrum of tumors associated with oncogenic Ras in humans often differs from those in mice either treated with carcinogens or engineered to sporadically express oncogenic Ras, suggesting that the mechanism of Ras transformation may be different in humans. Ras stimulates primarily three main classes of effector proteins, Rafs, PI3-kinase, and RalGEFs, with Raf generally being the most potent at transforming murine cells. Using oncogenic Ras mutants that activate single effectors as well as constitutively active effectors, we find that the RalGEF, and not the Raf or PI3-kinase pathway, is sufficient for Ras transformation in human cells. Thus, oncogenic Ras may transform murine and human cells by distinct mechanisms, and the RalGEF pathway-previously deemed to play a secondary role in Ras transformation-could represent a new target for anti-cancer therapy.