Analysis of endoplasmic reticulum trafficking signals by combinatorial screening in mammalian cells

Noa Zerangue, Michael J. Malan, Sharon R. Fried, Paul F. Dazin, Yuh Nung Jan, Lily Yeh Jan, Blanche Schwappach

    Research output: Contribution to journalArticlepeer-review

    Abstract

    To improve the accuracy of predicting membrane protein sorting signals, we developed a general methodology for defining trafficking signal consensus sequences in the environment of the living cell. Our approach uses retroviral gene transfer to create combinatorial expression libraries of trafficking signal variants in mammalian cells, flow cytometry to sort cells based on trafficking phenotype, and quantitative trafficking assays to measure the efficacy of individual signals. Using this strategy to analyze arginine- and lysine-based endoplasmic reticulum localization signals, we demonstrate that small changes in the local sequence context dramatically alter signal strength, generating a broad spectrum of trafficking phenotypes. Finally, using sequences from our screen, we found that the potency of di-lysine, but not di-arginine, mediated endoplasmic reticulum localization was correlated with the strength of interaction with α-COP.
    Original languageEnglish
    Pages (from-to)2431-2436
    Number of pages5
    JournalProceedings of the National Academy of Sciences of the United States of America
    Volume98
    Issue number5
    DOIs
    Publication statusPublished - 27 Feb 2001

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