Targeted expression of oncogenic K-ras in intestinal epithelium causes spontaneous tumorigenesis in mice

doi:10.1053/gast.2002.34786

Gastroenterology

Volume 123, Issue 2, August 2002, Pages 492-504

https://doi.org/10.1053/gast.2002.34786 Get rights and content

Abstract

Background & Aims: Ras oncoproteins are mutated in about 50% of human colorectal cancers, but their precise role in tumor initiation or progression is still unclear. Methods: This study presents transgenic mice that express K-ras^V12G, the most frequent oncogenic mutation in human tumors, under control of the murine villin promoter in epithelial cells of the large and small intestine. Results: More than 80% of the transgenic animals displayed single or multiple intestinal lesions, ranging from aberrant crypt foci (ACF) to invasive adenocarcinomas. Expression of K-ras^V12G caused activation of the MAP kinase cascade, and the tumors were frequently characterized by deregulated cellular proliferation. Unexpectedly, we obtained no evidence of inactivating mutations of the tumor suppressor gene Apc, the “gatekeeper” in colonic epithelial proliferation. However, spontaneous mutation of the tumor-suppressor gene p53, a frequent feature in the human disease, was found in 3 of 7 tumors that were tested. Conclusions: This animal model recapitulates the stages of tumor progression as well as a part of the genetic alterations found in human colorectal cancer. Furthermore, it indicates that activation of K-ras in concert with mutations in p53 may constitute a route to digestive tumor formation and growth, underlining the fact that the pathway to intestinal cancer is not necessarily a single road.

GASTROENTEROLOGY 2002;123:492-504

Section snippets

Establishment of the transgene and transgenic mouse lines

The vector pCEX-V3-V12-K-ras was generously provided by Dr. J. De Gunzburg (Inst. Curie, Paris, France). It contains the coding sequence of human K-ras4B with an activating glycine to valine mutation at codon 12 (V12G).²⁴ We modified the K-ras coding sequence by polymerase chain reaction (PCR) with sequence-specific primers that introduced a ScaI site at the 5' end and a NotI site at the 3' end (sense: 5'-TGCAAAAGTACTGAATATAAACTTGTG-3', antisense: 5'-ATTTGCGGCCGCTTTACATAATTACACACT-3'). PCR was

Generation of K-ras^V12G transgenic mice

A 9-kb regulatory region of the mouse villin gene was used to drive the selective expression of a K-ras^V12G transgene in epithelial cells of the large and small intestine. We have shown earlier that the villin regulatory region targets stable and homogeneous expression of transgenes all along the crypt-villus axis, in differentiated enterocytes as well as in the immature stem cells of the crypt.22, 23 Villin is an actin-binding protein that is expressed in immature, as well as in differentiated

Discussion

Earlier attempts to generate transgenic models for the role of oncogenic Ras in colorectal cancer have failed to produce neoplasms.11, 20 Expression of the transgene in postmitotic enterocytes was apparently not sufficient to induce cancer. We hypothesize that it is crucial to choose a promoter that targets expression of the transgene in the rapidly proliferating undifferentiated cell population, as it has been shown in a study focusing on the Wnt pathway.⁴⁸ These cells remain anchored in the

Acknowledgements

The authors thank Dr. Georges Gaudriault for the bacterial extracts containing recombinant RalGDS-GST, Dr. Philippe Vielh for allowing us to carry out the cytometry studies in his laboratory, Dr. Zofia Maciorowski for advice on cytometry, Dr. Jens Niewoehner for his help in constructing the K-ras^V12G vector, Dr. Mitsuhiro Tada for providing the mouse FASAY, Dr. Riccardo Fodde for generously providing the Apc1638N mouse strain, Reeta Holmila for assistance with the FASAY, and Louis Aussepe for

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^☆: Address requests for reprints to: Sylvie Robine, Ph.D., CNRS-UMR144, Institut Curie, 26 rue d'Ulm, 75248 Paris Cedex 05, France. e-mail: [email protected]; fax: (33) 1-4234-6377.

^☆☆: Supported by grants from the Association pour la Recherche contre le Cancer (ARC; to S.R.) and the Deutsche Forschungsgemeinschaft (DFG; to K.-P.J).

View full text

Basic–Alimentary TractTargeted expression of oncogenic K-ras in intestinal epithelium causes spontaneous tumorigenesis in mice☆,☆☆

Abstract

Section snippets

Establishment of the transgene and transgenic mouse lines

Generation of K-rasV12G transgenic mice

Discussion

Acknowledgements

Cell

Cell

Cell

Cell

Cell

Trends Cell Biol

Trends Cell Biol

J Biol Chem

Gastroenterology

Mutat Res

J Urol

Cell

J Biol Chem

J Biol Chem

Gastroenterology

Cell

Gastroenterology

Mol Cell

Mouse models for colorectal cancer

Oncogene

Multiple intestinal neoplasia caused by a mutation in the murine homolog of the APC gene

Science

A targeted chain-termination mutation in the mouse Apc gene results in multiple intestinal tumors

Proc Natl Acad Sci U S A

Colorectal carcinomas in mice lacking the catalytic subunit of PI(3)Kγ

Nature

Tumour biology. Weakening link to colorectal cancer?

Nature

Bi-transgenic mice reveal that K-rasVal12 augments a p53-independent apoptosis when small intestinal villus enterocytes reenter the cell cycle

J Cell Biol

Somatic activation of the K-ras oncogene causes early onset lung cancer in mice

Nature

Prevalence of ras gene mutations in human colorectal cancers

Nature

Detection of high incidence of K-ras oncogenes during human colon tumorigenesis

Nature

Activation of mitogen-activated protein kinase is necessary but not sufficient for proliferation of human thyroid epithelial cells induced by mutant Ras

Oncogene

PI-3-kinase is an essential anti-apoptotic effector in the proliferative response of primary human epithelial cells to mutant RAS

Oncogene

Transgenic mouse models that explore the multistep hypothesis of intestinal neoplasia

J Cell Biol

Frequent and characteristic K-ras activation in aberrant crypt foci of colon. Is there preference among K-ras mutants for malignant progression?

Cancer

Epithelial cell growth and differentiation. IV. Controlled spatiotemporal expression of transgenes: new tools to study normal and pathological states

Am J Physiol

Human colon carcinoma Ki-ras2 oncogene and its corresponding proto-oncogene

Mol Cell Biol

Suppression of villin expression by antisense RNA impairs brush border assembly in polarized epithelial intestinal cells

EMBO J

Cleavage of structural proteins during the assembly of the head of bacteriophage T4

Nature

Basic–Alimentary Tract
Targeted expression of oncogenic K-ras in intestinal epithelium causes spontaneous tumorigenesis in mice☆,☆☆

Generation of K-ras^V12G transgenic mice