Intracellular location of SNAP-25 in human neutrophils

Exocytosis plays an essential role in the physiological functions of human neutrophils. Although SNAP-25 is considered to play a key role in vesicle-membrane fusion, it has been detected almost exclusively in the neuronal system. Using different specific antibodies to SNAP-25, we have identified in the membrane fraction of resting human neutrophils an immunoreactive band with the same molecular mass observed in brain homogenates. Immunoblot analysis of subcellular fractions of neutrophils revealed that SNAP-25 protein was found in the granule membrane fraction, but not in the cytosolic and plasma membrane fractions. Granule localization for neutrophil SNAP-25 was further demonstrated by confocal and immunoelectron microscopy. Furthermore, SNAP-25 was mainly located in the morphologically defined neutrophil peroxidase-negative granules, which are mobilizable upon cell activation. In addition, the protein was specifically cleaved by botulinal neurotoxin A, as observed in brain homogenate. These findings reveal the presence of SNAP-25 in the granule membranes of human neutrophils.