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High-throughput retroviral tagging to identify components of specific signaling pathways in cancer

Nature Genetics volume 32, pages 153–159 (2002)Cite this article

A Corrigendum to this article was published on 01 November 2002

An Erratum to this article was published on 01 October 2002

Abstract

Genetic screens carried out in lower organisms such as yeast1, Drosophila melanogaster2 and Caenorhabditis elegans3 have revealed many signaling pathways. For example, components of the RAS signaling cascade were identified using a mutant eye phenotype in D. melanogaster as a readout2. Screening is usually based on enhancing or suppressing a phenotype by way of a known mutation in a particular signaling pathway. Such in vivo screens have been difficult to carry out in mammals, however, owing to their relatively long generation times and the limited number of animals that can be screened. Here we describe an in vivo mammalian genetic screen used to identify components of pathways contributing to oncogenic transformation. We applied retroviral insertional mutagenesis in Myc transgenic (EμMyc) mice lacking expression of Pim1 and Pim2 to search for genes that can substitute for Pim1 and Pim2 in lymphomagenesis. We determined the chromosomal positions of 477 retroviral insertion sites (RISs) derived from 38 tumors from EμMyc Pim1−/− Pim2−/− mice and 27 tumors from EμMyc control mice using the Ensembl and Celera annotated mouse genome databases. There were 52 sites occupied by proviruses in more than one tumor. These common insertion sites (CISs) are likely to contain genes contributing to tumorigenesis. Comparison of the RISs in tumors of Pim-null mice with the RISs in tumors of EμMyc control mice indicated that 10 of the 52 CISs belong to the Pim complementation group. In addition, we found that Pim3 is selectively activated in Pim-null tumor cells, which supports the validity of our approach.

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Figure 1: Retroviral tagging in lymphoma-prone EμMyc mice that are sensitized to activation of the Pim pathway.
Figure 2: Isolation of the genomic DNA sequences flanking the provirus using a splinkerette-based PCR approach.

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Notes

  1. NOTE: By error, several corrections were not made to proofs while preparing the manuscript for the press. This has now been corrected in the full-text of the article. The PDF of the article has not been corrected, but a copy of this amendment has been appended to the last page of the PDF. A printed erratum will be published in a forthcoming issue. The following corrections have been made: In the reference list, reference 25 (Losman et al.) has been inserted as reference 13. As a consequence, references 13-24 have been renumbered as 14-25. In the text, the following changes have been made: On page 154, in the second column, reference 16 has been placed at the end of the sentence "These observations underscore the selective advantage..." Reference 16 has also been removed from the first line on page 155. On page 155, in the second column, reference 17 has been placed at the end of the sentence "'Cold' and 'hot' spots for transposon insertions..." On page 157, in the first full paragraph, reference 17 has been changed to reference 18, reference 18 has been changed to reference 19, reference 19 has been changed to reference 20, and reference 25 has been changed to reference 21. On page 158, in the first full paragraph, references 21-23 have been changed to references 22,23.

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Acknowledgements

We wish to thank R. Regnerus for assistance in genotyping the mice;N. Bosnie, L. Rijswijk, A. Zwerver, T. Maidment, C. Spaans and F. van der Ahé for animal care; and J. Jonkers and R. van Amerongen for critical reading of the manuscript. This work was supported by the Dutch Cancer Society (H.M.) and the Leukemia Society of America (J.A.).

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  1. Division of Molecular Genetics and Centre of Biomedical Genetics, Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands

    Harald Mikkers, John Allen, Puck Knipscheer, Lieke Romeyn, Edwin Vink & Anton Berns

  2. Division of Radiotherapy, Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands

    Augustinus Hart

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Correspondence to Anton Berns.

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Mikkers, H., Allen, J., Knipscheer, P. et al. High-throughput retroviral tagging to identify components of specific signaling pathways in cancer. Nat Genet 32, 153–159 (2002). https://doi.org/10.1038/ng950

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