title: Canonical interaction of cyclin G associated kinase with adaptor protein 1 regulates lysosomal enzyme sorting.
abstract: The adaptor protein 1 (AP1) complex is a heterotetramer that participates in cargo sorting into clathrin-coated vesicles at the trans-Golgi network (TGN) and endosomes. The gamma subunit of AP1 possesses a C-terminal "ear" domain that recruits a cohort of accessory proteins through recognition of a shared canonical motif, PsiG[PDE][PsiLM] (where Psi is an aromatic residue). The physiological relevance of these ear-motif interactions, however, remains to be demonstrated. Here we report that the cyclin G-associated kinase (GAK) has two sequences fitting this motif, FGPL and FGEF, which mediate binding to the AP1-gamma-ear domain in vitro. Mutation of both gamma-ear-binding sequences or depletion of AP1-gamma by RNA interference (RNAi) decreases the association of GAK with the TGN in vivo. Depletion of GAK by RNAi impairs the sorting of the acid hydrolase, cathepsin D, to lysosomes. Importantly, expression of RNAi-resistant GAK restores the lysosomal sorting of cathepsin D in cells depleted of endogenous GAK, whereas expression of a similar construct bearing mutations in both gamma-ear-binding sequences fails to correct the sorting defect. Thus, interactions between the PsiG[PDE][PsiLM]-motif sequences in GAK and the AP1-gamma-ear domain are critical for the recruitment of GAK to the TGN and the function of GAK in lysosomal enzyme sorting.
authors.affiliation: Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, and Laboratory of Cell Biology, National Heart Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kametaka
authors.forename: Kengo
authors.initials: K
authors.lastname: Moriyama
authors.forename: Patricia V
authors.initials: PV
authors.lastname: Burgos
authors.forename: Evan
authors.initials: E
authors.lastname: Eisenberg
authors.forename: Lois E
authors.initials: LE
authors.lastname: Greene
authors.forename: Rafael
authors.initials: R
authors.lastname: Mattera
authors.forename: Juan S
authors.initials: JS
authors.lastname: Bonifacino
mesh_terms: D033961:Adaptor Protein Complex 1 / Q000378:metabolism*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002470:Cell Survival; D016384:Consensus Sequence; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008247:Lysosomes / Q000201:enzymology*; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D021601:trans-Golgi Network / Q000378:metabolism
chemical_list: D033961:Adaptor Protein Complex 1; D047908:Intracellular Signaling Peptides and Proteins; D010455:Peptides; C486325:GAK protein, human; D017346:Protein Serine-Threonine Kinases