• Lanford, Robert E (PI)

    Project: Research project

    Project Details


    The mechanism by which proteins are transported to the nucleus
    in eucaryotic cells will be examined. Elucidation of the
    mechanism(s) governing nuclear transport and nucleocytoplasmic
    interactions has broad implications to understanding the regulation
    of gene expression, oncogenic transformation, and the control of
    early embryonic development. The simiam virus 40 (SV40) large
    tumor (T) antigen offers an exceptional model for the analysis of
    nuclear transport; the T antigen polypeptide is well characterized,
    the role of T antigen in replication and oncogenic transformation
    is well established, the nuclear transport signal of T antigen is
    known, and the T antigen polypeptide is localized at both the
    nucleus and the cell surface. A system for the analysis of nuclear
    transport has been developed which utilizes microinjection of
    mammalian cells with synthetic peptide nuclear transport signals
    crosslinked to carrier proteins. A peptide homologous to the T
    antigen nuclear transport signal induces the nuclear transport of
    carrier proteins, but no transport occur when proteins are coupled
    to a synthetic peptide homologous to a nuclear-transport
    defective T antigen. The first specific aim will be to examine
    modified T antigen nuclear transport signal peptides, as well as
    signal peptides from other proteins for induction of nuclear
    transport. The flexibility of the nuclear transport machinery will
    be evaluated by measuring the kinetics of transport for modified
    peptides, and competitive microinjection experiments will
    determine whether different cellular factors are involved in
    recognition of dissimilar transport signals from other proteins.
    Second, the cellular protein that binds to the T antigen transport
    signal will be isolated and characterized. The signal peptide and
    anti-idiotypic antibodies that mimic the conformation of the
    signal peptide will be used to isolate cellular transport factors.
    The third specific aim will be to isolate and sequence the gene
    encoding the cellular factor that binds the T antigen signal
    sequence. The cloned gene will be used to analyze the
    conservation of the gene during eucaryotic evolution and to
    construct expression vectors. Fourth, a detailed immunological
    characterization of the ligand-receptor (signal sequence-transport
    factor) interaction will be performed. Finally, an attempt will be
    made to reconstitute an in vitro nuclear transport system using
    isolated nuclear transport factors.
    Effective start/end date7/1/845/31/92


    • National Institutes of Health: $174,726.00


    • Medicine(all)


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