A dileucine-like sorting signal directs transport into an AP-3-dependent, clathrin-independent pathway to the yeast vacuole.
Transport of yeast alkaline phosphatase (ALP) to the vacuole depends on the clathrin adaptor-like complex AP-3, but does not depend on proteins necessary for transport through pre-vacuolar endosomes. We have identified ALP sequences that direct sorting into the AP-3-dependent pathway using chimeric proteins containing residues from the ALP cytoplasmic domain fused to sequences from a Golgi-localized membrane protein, guanosine diphosphatase (GDPase). The full-length ALP cytoplasmic domain, or ALP amino acids 1-16 separated from the transmembrane domain by a spacer, directed GDPase chimeric proteins from the Golgi complex to the vacuole via the AP-3 pathway. Mutation of residues Leu13 and Val14 within the ALP cytoplasmic domain prevented AP-3-dependent vacuolar transport of both chimeric proteins and full-length ALP. This Leucine-Valine (LV)-based sorting signal targeted chimeric proteins and native ALP to the vacuole in cells lacking clathrin function. These results identify an LV-based sorting signal in the ALP cytoplasmic domain that directs transport into a clathrin-independent, AP-3-dependent pathway to the vacuole. The similarity of the ALP sorting signal to mammalian dileucine sorting motifs, and the evolutionary conservation of AP-3 subunits, suggests that dileucine-like signals constitute a core element for AP-3-dependent transport to lysosomal compartments in all eukaryotic cells.
Vowels
J J
JJ
Department of Biological Chemistry, UCLA School of Medicine, University of California, Los Angeles, Los Angeles, CA 90095, USA.
Payne
G S
GS
D033963:Adaptor Protein Complex 3; D000469:Alkaline Phosphatase / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D002966:Clathrin / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005838:Genotype; D006056:Golgi Apparatus / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D007930:Leucine; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011755:Pyrophosphatases / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D015398:Signal Transduction; D014157:Transcription Factors / Q000378:metabolism*; D014617:Vacuoles / Q000378:metabolism*; D014633:Valine
D033963:Adaptor Protein Complex 3; D002966:Clathrin; D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D000469:Alkaline Phosphatase; D011755:Pyrophosphatases; C010357:guanosine-diphosphatase; D007930:Leucine; D014633:Valine