Dominant effector genetics in mammalian cells

Abstract

We have expressed libraries of peptides in mammalian cells to select for trans-dominant effects on intracellular signaling systems. As an example—and to reveal pharmacologically relevant points in pathways that lead to Taxol resistance—we selected for peptide motifs that confer resistance to Taxol-induced cell death. Of several peptides selected, one, termed RGP8.5, was linked to upregulation of expression of the gene ABCB1 (also known as MDR1, for multiple drug resistance) in HeLa cells. Our data indicate that trans-dominant effector peptides can point to potential mechanisms by which signaling systems operate. Such tools may be useful in functional genomic analysis of signaling pathways in mammalian disease processes.

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Figure 1: Scheme for the peptide library screen.
Figure 2: Peptides conferring Taxol resistance to HeLa cells.
Figure 3: RGP8.5 peptide upregulates ABCB1 in HeLa cells.
Figure 4: ABCB1 expression conferring Taxol resistance to HeLa cells.
Figure 5: ABCB1 mRNA steady-state levels are increased by RGP8.5 peptide.
Figure 6: Association of the RGP8.5 peptide with PSMA7 and PSMA5 proteasome subunits.
Figure 7: Chemical inhibition of the proteasome also induces ABCB1 expression.

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Acknowledgements

We thank R. Armstrong and B. Fox for construction of the peptide library and retroviral vectors; C. Liao for critical reading of the manuscript; and R. Smith for suggestions on editing and clarity. G.P.N. is a recipient of the Burroughs Wellcome New Investigator Award in Pharmacology, is a Scholar of the Leukemia Society of America and is supported by the NIH.

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Correspondence to Xiang Xu or Garry P. Nolan.

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Xu, X., Leo, C., Jang, Y. et al. Dominant effector genetics in mammalian cells. Nat Genet 27, 23–29 (2001) doi:10.1038/83717

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