Targeting of P-selectin to two regulated secretory organelles in PC12 cells.
Targeting of P-selectin to the regulated secretory organelles (RSOs) of phaeochromocytoma PC12 cells has been investigated. By expressing from cDNA a chimera composed of HRP and P-selectin, and then following HRP activity through subcellular fractionation, we have discovered that P-selectin contains signals that target HRP to the synaptic-like microvesicles (SLMV) as well as the dense-core granules (DCGs) of these cells. Mutagenesis of the chimera followed by transient expression in PC12 cells shows that at least two different sequences within the carboxy-terminal cytoplasmic tail of P-selectin are necessary, but that neither is sufficient for trafficking to the SLMV. One of these sequences is centred on the 10 amino acids of the membrane-proximal C1 exon that is also implicated in lysosomal targeting. The other sequence needed for trafficking to the SLMV includes the last four amino acids of the protein. The same series of mutations have a different effect on DCG targeting, showing that traffic to the two different RSOs depends on different features within the cytoplasmic domain of P-selectin.
Norcott
J P
JP
Medical Research Council Laboratory for Molecular Cell Biology, University College London, United Kingdom.
Solari
R
R
Cutler
D F
DF
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003594:Cytoplasmic Granules / Q000378:metabolism*; D017931:DNA Primers; D004298:Dopamine / Q000378:metabolism; D004705:Endocytosis; D006801:Humans; D008969:Molecular Sequence Data; D015388:Organelles / Q000378:metabolism; D019007:P-Selectin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016716:PC12 Cells; D009195:Peroxidase / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D013347:Subcellular Fractions / Q000201:enzymology; D016708:Synaptophysin / Q000378:metabolism
D017931:DNA Primers; D019007:P-Selectin; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D016708:Synaptophysin; D009195:Peroxidase; D004298:Dopamine