Identification of mutant K-Ras-dependent phenotypes using a panel of isogenic cell lines

Steffan Vartanian, Carolyn Bentley, Matthew J. Brauer, Li Li, Senji Shirasawa, Takehiko Sasazuki, Jung Sik Kim, Pete Haverty, Eric Stawiski, Zora Modrusan, Todd Waldman, David Stokoe

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Abstract

To assess the consequences of endogenous mutant K-Ras, we analyzed the signaling and biological properties of a small panel of isogenic cell lines. These include the cancer cell lines DLD1, HCT116, and Hec1A, in which either the WT or mutant K-ras allele has been disrupted, and SW48 colorectal cancer cells and human mammary epithelial cells in which a single copy of mutant K-ras was introduced at its endogenous genomic locus. Wefind that single copy mutant K-Ras causes surprisingly modest activation of downstream signaling to ERK and Akt. In contrast, a negative feedback signaling loop to EGFR and N-Ras occurs in some, but not all, of these cell lines. Mutant K-Ras also had relatively minor effects on cell proliferation and cell migration but more dramatic effects on cell transformation as assessed by growth in soft agar. Surprisingly, knock-out of the wild type K-ras allele consistently increased growth in soft agar, suggesting tumor-suppressive properties of this gene under these conditions. Finally, we examined the effects of single copy mutant K-Ras on global gene expression. Although transcriptional programs triggered by mutant K-Ras were generally quite distinct in the different cell lines, there was a small number of genes that were consistently overexpressed, and these could be used to monitor K-Ras inhibition in a panel of human tumor cell lines. We conclude that there are conserved components of mutant K-Ras signaling and phenotypes but that many depend on cell context and environmental cues.

Original languageEnglish
Pages (from-to)2403-2413
Number of pages11
JournalJournal of Biological Chemistry
Volume288
Issue number4
DOIs
Publication statusPublished - Jan 25 2013

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Cells
Phenotype
Cell Line
Agar
Alleles
Tumors
Growth
Genes
Tumor Cell Line
Cell Movement
Cues
Colorectal Neoplasms
Neoplasms
Breast
Epithelial Cells
Cell Proliferation
Cell proliferation
Gene Expression
Gene expression
Chemical activation

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Vartanian, S., Bentley, C., Brauer, M. J., Li, L., Shirasawa, S., Sasazuki, T., ... Stokoe, D. (2013). Identification of mutant K-Ras-dependent phenotypes using a panel of isogenic cell lines. Journal of Biological Chemistry, 288(4), 2403-2413. https://doi.org/10.1074/jbc.M112.394130

Identification of mutant K-Ras-dependent phenotypes using a panel of isogenic cell lines. / Vartanian, Steffan; Bentley, Carolyn; Brauer, Matthew J.; Li, Li; Shirasawa, Senji; Sasazuki, Takehiko; Kim, Jung Sik; Haverty, Pete; Stawiski, Eric; Modrusan, Zora; Waldman, Todd; Stokoe, David.

In: Journal of Biological Chemistry, Vol. 288, No. 4, 25.01.2013, p. 2403-2413.

Research output: Contribution to journalArticle

Vartanian, S, Bentley, C, Brauer, MJ, Li, L, Shirasawa, S, Sasazuki, T, Kim, JS, Haverty, P, Stawiski, E, Modrusan, Z, Waldman, T & Stokoe, D 2013, 'Identification of mutant K-Ras-dependent phenotypes using a panel of isogenic cell lines', Journal of Biological Chemistry, vol. 288, no. 4, pp. 2403-2413. https://doi.org/10.1074/jbc.M112.394130
Vartanian S, Bentley C, Brauer MJ, Li L, Shirasawa S, Sasazuki T et al. Identification of mutant K-Ras-dependent phenotypes using a panel of isogenic cell lines. Journal of Biological Chemistry. 2013 Jan 25;288(4):2403-2413. https://doi.org/10.1074/jbc.M112.394130
Vartanian, Steffan ; Bentley, Carolyn ; Brauer, Matthew J. ; Li, Li ; Shirasawa, Senji ; Sasazuki, Takehiko ; Kim, Jung Sik ; Haverty, Pete ; Stawiski, Eric ; Modrusan, Zora ; Waldman, Todd ; Stokoe, David. / Identification of mutant K-Ras-dependent phenotypes using a panel of isogenic cell lines. In: Journal of Biological Chemistry. 2013 ; Vol. 288, No. 4. pp. 2403-2413.
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