Interaction of FK506-binding protein 13 with brefeldin A-inhibited guanine nucleotide-exchange protein 1 (BIG1): effects of FK506.

BIG1 and BIG2 are brefeldin A-inhibited guanine nucleotide-exchange proteins that activate ADP-ribosylation factors (ARFs), critical components of vesicular trafficking pathways. These proteins can exist in macromolecular complexes and move between Golgi membranes and cytosol. In the BIG1 molecule, a centrally located Sec7 domain is responsible for ARF activation, but functions ...
of other regions are largely unknown. Yeast two-hybrid screens of a human placenta cDNA library with BIG1 cDNA constructs revealed specific interaction of the N-terminal region (amino acids 1-331) with FK506-binding protein 13 (FKBP13). The association was confirmed by immunoprecipitation of both endogenous BIG1 and FKBP13 from Jurkat T cells with antibodies against either one. Binding of BIG1, BIG2, and ARF to cell membranes in vitro was increased by guanosine 5'-[gamma-thio]triphosphate, and further increases were induced by FK506. Incubation of Jurkat T cells with FK506 increased binding of BIG1, BIG2, and ARF to Golgi and other membranes in a time- and concentration-dependent manner, without effects on clathrin or gamma-adaptin binding. Binding of BIG1, BIG2, and ARF to membranes was also increased by L-732,531, an agonist structurally related to FK506, but was not increased by a related antagonist, L-685,818, nor by cyclosporin A or rapamycin. These findings are consistent with a role for FKBP13 and FK506 in vesicular trafficking, influencing ARF activity through their guanine nucleotide-exchange proteins.
Mesh Terms:
Adenosine Diphosphate, Blotting, Western, Brefeldin A, Cell Membrane, Clathrin, Cloning, Molecular, Cyclosporine, Dose-Response Relationship, Drug, Enzyme Inhibitors, GTP-Binding Proteins, Guanine Nucleotide Exchange Factors, Humans, Immunosuppressive Agents, Jurkat Cells, Precipitin Tests, Protein Binding, Sirolimus, Subcellular Fractions, Tacrolimus, Tacrolimus Binding Proteins, Temperature, Time Factors, Two-Hybrid System Techniques
Proc. Natl. Acad. Sci. U.S.A.
Date: Mar. 04, 2003
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