title: Differential binding of arfaptin 2/POR1 to ADP-ribosylation factors and Rac1.
abstract: We first identified arfaptin as a protein that bound to GTP-ARFs (especially ARF1). However, a second group reported that POR1, a truncated form of arfaptin, bound to GTP-Rac1. Therefore, we examined the possibility that arfaptin 2/POR1 was a common downstream effector for both ARF1 and Rac1. In this study, we found that constitutively active Rac1 or GTP-Rac1 showed negligible or no binding to arfaptin 2/POR1 in a yeast two-hybrid assay or a GST pull-down assay. However, wild-type or dominant negative Rac1 or Rac1 liganded to GDP showed strong binding. In contrast, constitutively active ARFs1, 5, and 6 showed binding, whereas the wild-type and dominant negative forms did not. Furthermore, the GTP-liganded ARFs bound arfaptin 2, whereas the GDP-bound forms showed little or no binding. Based on these observations, we suggest that arfaptin 2/POR1 is a target protein for GTP-ARFs and for GDP-Rac1, and that it may be involved in interactions between the Rac and ARF signaling pathways.
authors.affiliation: Howard Hughes Medical Institute, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0295, USA.
authors.forename: O H
authors.initials: OH
authors.lastname: Shin
authors.forename: J H
authors.initials: JH
authors.lastname: Exton
mesh_terms: D020727:ADP-Ribosylation Factors / Q000235:genetics / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D016466:CHO Cells; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D006224:Cricetinae; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression; D005799:Genes, Dominant; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006153:Guanosine Diphosphate / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011485:Protein Binding / Q000187:drug effects / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D020798:Two-Hybrid System Techniques; D020830:rac1 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: C101158:ARFIP2 protein, human; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D005982:Glutathione Transferase; D020727:ADP-Ribosylation Factors; D020830:rac1 GTP-Binding Protein