Vol. 12, Issue 3, 663-673, March 2001

Molecular Cloning and Characterization of Phocein, a Protein Found from the Golgi Complex to Dendritic Spines

Gilbert Baillat,^* Abdelaziz Moqrich,^* Francis Castets,^* Agnès Baude, Yannick Bailly, Alexandre Benmerah,^§ and Ariane Monneron^*

 ^*FRE Centre National de la Recherche Scientifique 9041 and  FRE Centre National de la Recherche Scientifique 9024, Centre National de la Recherche Scientifique, 13009 Marseille, France;  FRE Centre National de la Recherche Scientifique 2180, 67084 Strasbourg, France; and  ^§Institut National de la Santé et de la Recherche Médicale E9925, Faculté Necker-Enfants Malades, 75730 Paris, France

Phocein is a widely expressed, highly conserved intracellular protein of 225 amino acids, the sequence of which has limited homology to the  subunits from clathrin adaptor complexes and contains an additional stretch bearing a putative SH3-binding domain. This sequence is evolutionarily very conserved (80% identity between Drosophila melanogaster and human). Phocein was discovered by a yeast two-hybrid screen using striatin as a bait. Striatin, SG2NA, and zinedin, the three mammalian members of the striatin family, are multimodular, WD-repeat, and calmodulin-binding proteins. The interaction of phocein with striatin, SG2NA, and zinedin was validated in vitro by coimmunoprecipitation and pull-down experiments. Fractionation of brain and HeLa cells showed that phocein is associated with membranes, as well as present in the cytosol where it behaves as a protein complex. The molecular interaction between SG2NA and phocein was confirmed by their in vivo colocalization, as observed in HeLa cells where antibodies directed against either phocein or SG2NA immunostained the Golgi complex. A 2-min brefeldin A treatment of HeLa cells induced the redistribution of both proteins. Immunocytochemical studies of adult rat brain sections showed that phocein reactivity, present in many types of neurons, is strictly somato-dendritic and extends down to spines, just as do striatin and SG2NA.