AP-4, a Novel Protein Complex Related to Clathrin Adaptors

Abstract

Here we report the identification and characterization of AP-4, a novel protein complex related to the heterotetrameric AP-1, AP-2, and AP-3 adaptors that mediate protein sorting in the endocytic and late secretory pathways. The key to the identification of this complex was the cloning and sequencing of two widely expressed, mammalian cDNAs encoding new homologs of the adaptor ␤ and subunits named ␤4 and 4, respectively. An antibody to ␤4 recognized in human cells an ϳ83-kDa polypeptide that exists in both soluble and membrane-associated forms. Gel filtration, sedimentation velocity, and immunoprecipitation experiments revealed that ␤4 is a component of a multisubunit complex (AP-4) that also contains the 4 polypeptide and two additional adaptor subunit homologs named 4 (-ARP2) and ⑀. Immunofluorescence analyses showed that AP-4 is associated with the trans-Golgi network or an adjacent structure and that this association is sensitive to the drug brefeldin A. We propose that, like the related AP-1, AP-2, and AP-3 complexes, AP-4 plays a role in signal-mediated trafficking of integral membrane proteins in mammalian cells.

Genomic Structure of the Mouse Ap3b1 Gene in Normal and Pearl Mice

“…No curative therapies exist. HPS is a genetically heterogenous disease and is found in diverse populations worldwide (Hazelwood et al, 1997;Oh et al, 1998;Shotelersuk and Gahl, 1998;Spritz, 1999;Dell'Angelica et al, 1999). This is consistent with the existence of at least 14 genetically distinct mouse pigment mutants that exhibit phenotypes similar to those of HPS patients (Swank et al, 1998;Swank et al, in press).…”

“…Recently, HPS patients with compound heterozygous mutations in the HPS2 gene were reported (Dell'Angelica et al, 1999). A method to screen additional HPS patients for mutations in this gene would be useful.…”

“…The pearl gene, now designated Ap3b1 , and its human orthologue ADTB3A (Dell'Angelica et al, 1999) encode the ␤3A subunit of the AP-3 adaptor complex, which is involved in sorting specific membrane proteins and facilitating carrier vesicle formation at the trans-Golgi network Simpson et al, 1997). These vesicles are then targeted to their specific subcellular destinations, including lysosomes and presumably platelet dense granules and melanosomes.…”

“…Two pearl (pe) mutations, a 793-bp duplication in pearl and a 107-bp deletion in pearl-8J and compound mutations in HPS patients (Dell'Angelica et al, 1999), have been reported. These mutations lead to the depletion of the ␤3A subunit, which in turn results in a decrease or an abolition of other subunits of the AP-3 complex .…”

“…These mutations lead to the depletion of the ␤3A subunit, which in turn results in a decrease or an abolition of other subunits of the AP-3 complex . Mislocalization of the entire AP-3 complex and loss of its function with subsequent defects in all three above-mentioned organelles ensue (Dell'Angelica et al, 1999;Zhen et al, 1999). The importance of the AP-3 complex in SPD and in the biosynthesis/function of subcellular organelles is further confirmed by organellar alterations in another HPS mouse model, mocha (Kantheti et al, 1998), in Drosophila pigmentation mutations garnet , carmine , and ruby (Kretzschmar et al, 2000), and in yeast (Cowles et al, 1997;Stepp et al, 1997).…”