Phosphatidylinositol-4-kinase type II alpha contains an AP-3-sorting motif and a kinase domain that are both required for endosome traffic.
The adaptor complex 3 (AP-3) targets membrane proteins from endosomes to lysosomes, lysosome-related organelles and synaptic vesicles. Phosphatidylinositol-4-kinase type II alpha (PI4KIIalpha) is one of several proteins possessing catalytic domains that regulate AP-3-dependent sorting. Here we present evidence that PI4KIIalpha uniquely behaves both as a membrane protein cargo as well as an enzymatic regulator of adaptor function. In fact, AP-3 and PI4KIIalpha form a complex that requires a dileucine-sorting motif present in PI4KIIalpha. Mutagenesis of either the PI4KIIalpha-sorting motif or its kinase-active site indicates that both are necessary to interact with AP-3 and properly localize PI4KIIalpha to LAMP-1-positive endosomes. Similarly, both the kinase activity and the sorting signal present in PI4KIIalpha are necessary to rescue endosomal PI4KIIalpha siRNA-induced mutant phenotypes. We propose a mechanism whereby adaptors use canonical sorting motifs to selectively recruit a regulatory enzymatic activity to restricted membrane domains.
Craige
Branch
B
Graduate Program in Biochemistry, Cell, and Developmental Biology, Emory University, Atlanta, GA 30322, USA.
Salazar
Gloria
G
Faundez
Victor
V
D020816:Amino Acid Motifs; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D020134:Catalytic Domain / Q000235:genetics; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011992:Endosomes / Q000378:metabolism*; D006801:Humans; D051907:Lysosomal Membrane Proteins / Q000378:metabolism; D015778:Minor Histocompatibility Antigens; D046912:Multiprotein Complexes; D016297:Mutagenesis, Site-Directed; D016716:PC12 Cells; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D052118:Lysosomal-Associated Membrane Protein 1
D004268:DNA-Binding Proteins; D051907:Lysosomal Membrane Proteins; D015778:Minor Histocompatibility Antigens; D046912:Multiprotein Complexes; D017853:Phosphotransferases (Alcohol Group Acceptor); D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C489116:LAMP1 protein, human; D052118:Lysosomal-Associated Membrane Protein 1; C060759:enhancer-binding protein AP-3; C402182:phosphatidylinositol phosphate 4-kinase