## 4 GOLPH3 Interacts with COPI to Recruit Glycostransferases to Golgi Compartments  
Golgi phosphoprotein 3 (GOLPH3), a highly conserved protein from yeast to humans, has been characterized as a Phosphatidylinositol 4-phosphate [PI(4)P] effector that regulates Golgi trafficking [58]. Identified in proteomic-based studies of the Golgi [59; 60], GOLPH3 localizes to the _trans_-Golgi via the direct interaction with PI(4)P, mediated by its unique C-terminal GPP34 domain [61; 62]. Localization of GOLPH3 protein to the Golgi is required to maintain Golgi architecture in human cells and _Drosophila melanogaster_[61; 63]. Much evidence indicates that GOLPH3 is an oncogene, that is overexpressed in several solid tumors such as lung cancer, breast cancer, colon cancer and melanoma [58; 64]. However, the molecular mechanisms underlying the oncogenic properties of GOLPH3 are poorly understood. It has been proposed that GOLPH3 might contribute to cellular transformation by affecting the glycosylation of key cancer relevant glycoproteins [58]. Experimental data from human cells and in model organisms indicate that GOLPH3 plays a fundamental role for retrograde intra-Golgi trafficking of protein glycosyltransferases ([58]; Figure 3). Glycosyltransferases are all type II integral membrane proteins containing of a short cytosolically exposed \(N\)-terminal region, a single membrane-spanning domain and a luminal enzymatic domain [5]. The cytoplasmic tails of Golgi glycosyltransferase lacked the canonical COPI-binding sorting signals. The role of GOLPH3 in enzyme recycling was initially demonstrated for the yeast homologue Vps74p which was shown to bind the \(N\)-terminal tails of a set of manosyltransferases as well as the COPI coatomer, thereby anchoring these enzymes at the Golgi [67; 68]. Mutations in the PI(4)P-binding pocket of Vps74p caused loss of Vps74p-membrane association in yeast cells and mislocalization of manosyltransferases [69]. Both GOLPH3 and Vps74p form oligomers and tetramerization of Vps74p was essential to bind to a pentameric sequence motif at the cytoplasmic tail of a subset of Golgi glycosyltransferase in _in vitro_ assays [70]. A cluster of evolutionarily conserved arginine residues at the \(N\)-terminus of GOLPH3 proteins was shown to mediate coatomer binding [70]. Vps74p-coatomer binding was required for retention of glycosyltransferase to the Golgi, but not for Vps74p association with the Golgi membrane which was instead dependent on the oligomerization status and the PI(4)P binding capabilities [70]. Vps74p recognizes a (F/L)(L/I/V)XX(R/K) motif in the cytoplasmic tail of numerous yeast mannosyltransferases raising the possibility that it might function as an adaptor for incorporation of glycosyltransferases in the COPI vesicles. A sequence similar to the (F/L)(L/I/V)XX(R/K) motif, was also found in the cytoplasmic tail (CT) of Core 2 \(N\)-acetylglucosaminyltransferase 1 (C2 GnT1; [71]). C2 GnT1 plays a crucial role in the synthesis of core 2-associated sialyl Lewis x  
Figure 3: Proposed models for roles of GORAB and GOLPH3 in COPI-mediated trafficking. (Upper panel) The diagram depicts a model for GOLPH3 role in \(N\)-glycosylation based on the results of [PERSON] and coauthors [65]. GOLPH3 protein binds to the coatomer as well as to a set of glycosyltransferases, such as C2 GnT and SiTaT as well as the coatomer, thereby mediating incorporation of these enzymes into COPI coated vesicles. (Lower panel) The diagram depicts a model for GORAB role in COPI vesicle release based on the results of [PERSON] and coauthors [66] GORAB self-associates in oligomers that form discrete domains at the _trans_-Golgi membrane. GORAB oligomers recruit Scyl1 protein driving Arf1-GTP accumulation in the GORAB-enriched domains. High levels of Scyl1 and Arf1-GTP proteins lead to the selective COPI loading. In turn, coatomer assembly leads to vesicle budding and cargo incorporation while GORAB/Scyl1 complex stabilizes the bud neck during vesicle formation.