title: Apicoplast targeting of a Toxoplasma gondii transmembrane protein requires a cytosolic tyrosine-based motif.
abstract: Toxoplasma gondii, like most apicomplexan parasites, possesses an essential relict chloroplast, the apicoplast. Several apicoplast membrane proteins lack the bipartite targeting sequences of luminal proteins. Vesicles bearing these membrane proteins are detected during apicoplast enlargement, but the means of cargo selection remains obscure. We used a combination of deletion mutagenesis, point mutations and protein chimeras to identify a short motif prior to the first transmembrane domain of the T. gondii apicoplast phosphate transporter 1 (APT1) that is necessary for apicoplast trafficking. Tyrosine 16 was essential for proper localization; any substitution resulted in misdirection of APT1 to the Golgi body. Glycine 17 was also important, with significant Golgi body accumulation in the alanine mutant. Separation of at least eight amino acids from the transmembrane domain was required for full motif function. Similarly placed YG motifs are present in apicomplexan APT1 orthologs and the corresponding N-terminal domain from Plasmodium vivax was able to route T. gondii APT1 to the apicoplast. Differential permeabilization showed that both the N- and C-termini of APT1 are exposed to the cytosol. We propose that this YG motif facilitates APT1 trafficking via interactions that occur on the cytosolic face of nascent vesicles destined for the apicoplast.
authors.affiliation: Seattle Biomedical Research Institute, 307 Westlake Ave N, Seattle, WA 98109-5219, USA.
authors.forename: Amy E
authors.initials: AE
authors.lastname: DeRocher
authors.forename: Anuradha
authors.initials: A
authors.lastname: Karnataki
authors.forename: Pashmi
authors.initials: P
authors.lastname: Vaney
authors.forename: Marilyn
authors.initials: M
authors.lastname: Parsons
mesh_terms: D000409:Alanine / Q000737:chemistry; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D017353:Gene Deletion; D005998:Glycine / Q000737:chemistry; D006056:Golgi Apparatus / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism*; D008856:Microscopy, Fluorescence / Q000379:methods; D008969:Molecular Sequence Data; D010966:Plasmodium vivax / Q000378:metabolism; D018087:Plastids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D014122:Toxoplasma / Q000378:metabolism*; D014443:Tyrosine / Q000737:chemistry*
chemical_list: D008565:Membrane Proteins; D014443:Tyrosine; D000409:Alanine; D005998:Glycine