TARGETING OF P-SELECTIN TO 2 REGULATED SECRETORY ORGANELLES IN PC12 CELLS

Targeting of P-selectin to the regulated secretory organelles (RSOs) o f phaeochromocytoma PC12 cells has been investigated. By expressing fr om cDNA a chimera composed of HRP and P-selectin, and then following H RP activity through subcellular fractionation, we have discovered that P-selectin contains signals that target HRP to the synaptic-like micr ovesicles (SLMV) as well as the dense-core granules (DCGs) of these ce lls. Mutagenesis of the chimera followed by transient expression in PC 12 cells shows that at least two different sequences within the carbox y-terminal cytoplasmic tail of P-selectin are necessary, but that neit her 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 t argeting, showing that traffic to the two different RSOs depends on di fferent features within the cytoplasmic domain of P-selectin.