title: The novel Drosophila lysosomal enzyme receptor protein mediates lysosomal sorting in mammalian cells and binds mammalian and Drosophila GGA adaptors.
abstract: Biogenesis of lysosomes depends in mammalian cells on the specific recognition and targeting of mannose 6-phosphate-containing lysosomal enzymes by two mannose 6-phosphate receptors (MPR46, MPR300), key components of the extensively studied receptor-mediated lysosomal sorting system in complex metazoans. In contrast, the biogenesis of lysosomes is poorly investigated in the less complex metazoan Drosophila melanogaster. We identified the novel type I transmembrane protein lysosomal enzyme receptor protein (LERP) with partial homology to the mammalian MPR300 encoded by Drosophila gene CG31072. LERP contains 5 lumenal repeats that share homology to the 15 lumenal repeats found in all identified MPR300. Four of the repeats display the P-lectin type pattern of conserved cysteine residues. However, the arginine residues identified to be essential for mannose 6-phosphate binding are not conserved. The recombinant LERP protein was expressed in mammalian cells and displayed an intracellular localization pattern similar to the mammalian MPR300. The LERP cytoplasmic domain shows highly conserved interactions with Drosophila and mammalian GGA adaptors known to mediate Golgi-endosome traffic of MPRs and other transmembrane cargo. Moreover, LERP rescues missorting of soluble lysosomal enzymes in MPR-deficient cells, giving strong evidence for a function that is equivalent to the mammalian counterpart. However, unlike the mammalian MPRs, LERP did not bind to the multimeric mannose 6-phosphate ligand phosphomannan. Thus ligand recognition by LERP does not depend on mannose 6-phosphate but may depend on a common feature present in mammalian lysosomal enzymes. Our data establish a potential important role for LERP in biogenesis of Drosophila lysosomes and suggest a GGA function also in the receptor-mediated lysosomal transport system in the fruit fly.
authors.affiliation: Institut für Physiologische Chemie und Pathobiochemie, Universitaets-Klinikum-Muenster, Waldeyerstr. 15, D-48149 Muenster, Germany. dennes@uni-muenster.de
authors.forename: André
authors.initials: A
authors.lastname: Dennes
authors.forename: Christoph
authors.initials: C
authors.lastname: Cromme
authors.forename: Koduru
authors.initials: K
authors.lastname: Suresh
authors.forename: Nadimpalli Siva
authors.initials: NS
authors.lastname: Kumar
authors.forename: Johannes A
authors.initials: JA
authors.lastname: Eble
authors.forename: Anke
authors.initials: A
authors.lastname: Hahnenkamp
authors.forename: Regina
authors.initials: R
authors.lastname: Pohlmann
mesh_terms: D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D033942:Adaptor Proteins, Vesicular Transport / Q000378:metabolism*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000737:chemistry; D002402:Cathepsin D / Q000737:chemistry; D056668:Cathepsin L; D002403:Cathepsins / Q000737:chemistry; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002845:Chromatography; D017124:Conserved Sequence; D003546:Cysteine Endopeptidases / Q000737:chemistry; D003593:Cytoplasm / Q000378:metabolism; D004331:Drosophila melanogaster; D005347:Fibroblasts / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D047468:Immunoprecipitation; D037102:Lectins / Q000378:metabolism; D008024:Ligands; D008247:Lysosomes / Q000378:metabolism*; D008351:Mannans / Q000737:chemistry; D008360:Mannosephosphates / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D011485:Protein Binding; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D012333:RNA, Messenger / Q000378:metabolism; D017527:Receptor, IGF Type 2; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry* / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014162:Transfection
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; C407306:GGA adaptor proteins; D037102:Lectins; D008024:Ligands; D008351:Mannans; D008360:Mannosephosphates; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D017527:Receptor, IGF Type 2; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C034304:cation-dependent mannose-6-phosphate receptor; C027693:mannose-6-phosphate; C005437:phosphomannan; D001120:Arginine; D006026:Glycoside Hydrolases; D002403:Cathepsins; D003546:Cysteine Endopeptidases; D056668:Cathepsin L; C534278:Ctsl protein, mouse; D002402:Cathepsin D; D020727:ADP-Ribosylation Factors