title: Sorting of encystation-specific cysteine protease to lysosome-like peripheral vacuoles in Giardia lamblia requires a conserved tyrosine-based motif.
abstract: Encystation-specific cysteine protease (ESCP) was the first membrane-associated protein described to be part of the lysosome-like peripheral vacuoles in the intestinal parasite Giardia lamblia. ESCP is homologous to cathepsin C enzymes of higher eukaryotes, but is distinguished from other lysosomal cysteine proteases because it possesses a transmembrane domain and a short cytoplasmic tail. Tyrosine-based motifs within tails of membrane proteins are known to participate in endosomal/lysosomal protein sorting in higher eukaryotes. In this study, we show that a YRPI motif within the ESCP cytoplasmic tail is necessary and sufficient to mediate ESCP sorting to peripheral vacuoles in Giardia. Deletion and point mutation analysis demonstrated that the tyrosine residue is critical for ESCP sorting, whereas amino acids located at the Y+1 (Arg), Y+2 (Pro), and Y+3 (Ile) positions show minimal effect. Loss of the motif resulted in surface localization, whereas addition of the motif to a variant-specific surface protein resulted in lysosomal localization. Although Giardia trophozoites lack a morphologically discernible Golgi apparatus, our findings indicate that this parasite directs proteins to the lysosomes using a conserved sorting signal similar to that used by yeast and mammalian cells. Because Giardia is one of the earliest branching protist, these results demonstrate that sorting motifs for specific protein traffic developed very early during eukaryotic evolution.
authors.affiliation: Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA. mtouz@niaid.nih.gov
authors.forename: Maria C
authors.initials: MC
authors.lastname: Touz
authors.forename: Hugo D
authors.initials: HD
authors.lastname: Lujan
authors.forename: Stanley F
authors.initials: SF
authors.lastname: Hayes
authors.forename: Theodore E
authors.initials: TE
authors.lastname: Nash
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017124:Conserved Sequence; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D014644:Genetic Variation; D016829:Giardia lamblia / Q000201:enzymology* / Q000502:physiology / Q000648:ultrastructure; D008247:Lysosomes / Q000201:enzymology* / Q000648:ultrastructure; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014162:Transfection; D014443:Tyrosine; D014617:Vacuoles / Q000201:enzymology* / Q000648:ultrastructure
chemical_list: D011994:Recombinant Proteins; D014443:Tyrosine; D003546:Cysteine Endopeptidases