Localization of TGN38 to the trans-Golgi network: involvement of a cytoplasmic tyrosine-containing sequence.
Protein localization to the TGN was investigated by examining the subcellular distribution of chimeric proteins in which the cytoplasmic and/or transmembrane domains of the TGN protein, TGN38, were substituted for the analogous domains of the plasma membrane protein, Tac. Using immunofluorescence and immunoelectron microscopy, the COOH-terminal cytoplasmic domain of TGN38 was found to be sufficient for localization of the chimeric proteins to the TGN. Deletion analysis identified an 11-amino acid segment containing the critical sequence, YQRL, as being sufficient for TGN localization. TGN localization was abrogated by mutation of the tyrosine or leucine residues in this sequence to alanine, or of the arginine residue to aspartate. In addition to specifying TGN localization, the 11-amino acid segment was active as an internalization signal, although the property of internalization alone was insufficient to confer TGN localization. Overexpression of chimeric proteins containing TGN localization determinants resulted in their detection at the plasma membrane and in intracellular vesicles, and abolished detection of endogenous TGN38. These results suggest that discrete cytoplasmic determinants can mediate protein localization to the TGN, and reveal a novel role for tyrosine-based motifs in this process.
Humphrey
J S
JS
Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892.
Peters
P J
PJ
Yuan
L C
LC
Bonifacino
J S
JS
D000595:Amino Acid Sequence; D000818:Animals; D002451:Cell Compartmentation; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006023:Glycoproteins; D006056:Golgi Apparatus / Q000737:chemistry*; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D014162:Transfection
D006023:Glycoproteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C513045:Tgoln2 protein, rat