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Overexpressed vs mutated Kras in murine fibroblasts: a molecular phenotyping study
M Horsch1, C V Recktenwald, S Schädler
1Helmholtz Zentrum München-German Research Center for Environmental Health (GmbH), Institute of Experimental Genetics, Ingolstädter Landstrasse 1, 85764 Neuherberg, Germany.
Abstract:
Ras acts in signalling pathways regulating the activity of multiple cellular functions including cell proliferation, differentiation, and apoptosis. Amino-acid exchanges at position 12, 13, or 61 of the Kras gene convert the proto-oncogene into an activated oncogene. Until now, a direct comparison of genome-wide expression profiling studies of Kras overexpression and different Kras mutant forms in a single assay system has not been carried out. In our study, we focused on the direct comparison of global gene expression effects caused by mutations in codon 12 or 13 of the Kras gene and Kras overexpression in murine fibroblasts. We determined Kras cellular mRNA, Ras protein and activated Ras protein levels. Further, we compared our data to the proteome analysis of the same transfected cell lines. Both overexpression and mutations of Kras lead to common altered gene expression patterns. Only two genes, Lox and Col1a1, were reversely regulated in the Kras transfectants. They may contribute to the higher aggressiveness of the Kras codon 12 mutation in tumour progression. The functional annotation of differentially expressed genes revealed a high frequency of proteins involved in tumour growth and angiogenesis. These data further support the important role of these genes in tumour-associated angiogenesis.
Insights
Ras gene mutations and overexpression significantly alter cellular gene expression, impacting tumor growth and angiogenesis. Specific Kras mutations may drive tumor aggressiveness, highlighting their oncogenic role.
Area of Science:
- Molecular Biology
- Oncology
- Genetics
Background:
- Ras proteins are key regulators of cellular functions like proliferation and apoptosis.
- Mutations in the Kras gene, particularly at codons 12, 13, or 61, can activate it as an oncogene.
- Previous studies have not directly compared the genome-wide expression profiles of Kras overexpression versus its mutant forms within a single experimental system.
Purpose of the Study:
- To directly compare the global gene expression effects of Kras gene mutations (codon 12 or 13) and Kras overexpression in murine fibroblasts.
- To analyze Kras mRNA, Ras protein, and activated Ras protein levels.
- To correlate gene expression data with proteome analysis from the same cell lines.
Main Methods:
- Gene expression profiling of murine fibroblasts transfected with Kras overexpression or Kras mutants (codon 12 or 13).
- Measurement of Kras mRNA, Ras protein, and activated Ras protein levels.
- Proteome analysis of the transfected cell lines.
- Functional annotation of differentially expressed genes.
Main Results:
- Both Kras overexpression and mutations induced common alterations in gene expression patterns.
- Lox and Col1a1 genes showed reverse regulation in Kras transfectants, potentially linked to tumor aggressiveness.
- Functional analysis identified a high prevalence of differentially expressed genes involved in tumor growth and angiogenesis.
Conclusions:
- Kras alterations (overexpression and mutations) significantly impact global gene expression, affecting pathways crucial for cancer progression.
- The reverse regulation of Lox and Col1a1 by Kras mutations suggests a role in enhanced tumor aggressiveness.
- The findings underscore the critical involvement of Kras-regulated genes in tumor-associated angiogenesis and growth.
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