Abstract
p75NTR is a low-affinity nerve growth factor receptor, which promotes cell proliferation as a positive modulator of high-affinity receptor TrkA, as well as binds with cell ligands to induce apoptosis and mediate death signals. To analyze the regulatory mechanisms of p75NTR, the present study utilized a new membrane yeast two-hybrid system to screen a human fetal brain cDNA library. Results identified BFAR, a novel protein that interacts with p75NTR. Interaction specificity was verified by membrane yeast two-hybrid co-transformation assays, in vitro GST pull-down assays, and in vitro co-immunoprecipitation assays. The fluorescent subcellular localization assay revealed that the two proteins co-localized within the cytoplasm. BFAR overexpression in PC-12 and HEK293T cells inhibited the NFκB and JNK signaling pathway, as determined with the luciferase test. Co-transfected p75NTR and BFAR in HEK293T or PC-12 cells, respectively, increased the percentage of cells in the G2/M phase, decreased the number of S-phase cells, and did not change the number of G0/G1-phase cells.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adaptor Proteins, Signal Transducing / genetics
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Adaptor Proteins, Signal Transducing / metabolism*
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Animals
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Apoptosis Regulatory Proteins / genetics
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Apoptosis Regulatory Proteins / metabolism*
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Brain / embryology
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Brain / metabolism
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Cell Cycle / genetics
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Cell Cycle / physiology
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Cytoplasm / metabolism*
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Flow Cytometry
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Gene Library
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Green Fluorescent Proteins / genetics
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Green Fluorescent Proteins / metabolism
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HEK293 Cells
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HeLa Cells
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Humans
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Immunoprecipitation
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Membrane Proteins / genetics
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Membrane Proteins / metabolism*
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Microscopy, Fluorescence
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Nerve Tissue Proteins / genetics
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Nerve Tissue Proteins / metabolism*
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PC12 Cells
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Protein Binding
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Rats
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Receptor, Nerve Growth Factor / genetics
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Receptor, Nerve Growth Factor / metabolism*
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Receptors, Nerve Growth Factor / genetics
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Receptors, Nerve Growth Factor / metabolism*
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Signal Transduction / genetics
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Signal Transduction / physiology*
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Transfection
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Two-Hybrid System Techniques
Substances
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Adaptor Proteins, Signal Transducing
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Apoptosis Regulatory Proteins
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BFAR protein, human
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Membrane Proteins
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NGFR protein, human
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Nerve Tissue Proteins
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Receptor, Nerve Growth Factor
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Receptors, Nerve Growth Factor
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Green Fluorescent Proteins