PERICENTRIOLAR TARGETING OF GDP-DISSOCIATION INHIBITOR ISOFORM-2

Cellular mechanisms for regulating membrane movements appear to involv e small GTPases of the Rab subfamily. Binding of GDP-bound Rab protein s to donor membranes and their release from target membranes appear to be regulated by GDP-dissociation inhibitor (GDI) protein isoforms. Pr evious work showed strikingly higher Levels of GDI-2 than GDI-1 fracti onate with total membranes of cultured cells and are visualized in the perinuclear region in 3T3-L1 adipocytes. Here we report that GDI-2-co ntaining structural elements are concentrated predominantly in the per icentriolar area in interphase CHO-T cells and differentiated 3T3-L1 a dipocytes based on colocalization of GDI-2 and the centrosomal marker pericentrin. This finding is documented by both immunofluorescence and immunoelectron microscopy. Expressed c-Myc-tagged GDI-2 in transfecte d COS-7 cells targets to the same region. During mitotic resolution of the centrosome into two identifiable foci in CHO-T cells, GDI-2 conta ining structures remain intact and also resolve into two regions surro unding the centrosome. Dissociation of pericentriolar GDI-2 from the G olgi markers P-COP and lectin receptors was apparent upon brefeldin A treatment of 3T3-L1 adipocytes or CHO-T cells. The integrity of perice ntriolar GDI-2-binding elements was not disrupted by either brief Trit on X-100 extraction or microtubule cytoskeletal disassembly, achieved with nocodazole. These data demonstrate the presence of highly ordered , detergent-resistant GDI-2-specific structural elements around the ce ntrosome and indicate functional differences for the GDI-1 and GDI-2 p rotein isoforms. The results suggest the presence of selective GDI-2 a ccepters in this region and a possible role of pericentriolar GDI-2 in membrane recycling.