Identification of glyceraldehyde-3-phosphate dehydrogenase as a Ca2+-dependent fusogen in human neutrophil cytosol.

Abstract

The membrane fusion events observed during neutrophil degranulation are important aspects of the immunoregulatory system. In an attempt to understand the regulation of granule-plasma membrane fusion, we have begun characterizing human neutrophil cytosol for fusion activity, finding that 50% of the fusogenic activity could be attributed to members of the annexin family of proteins. The major non-annexin fusion activity (25% of the total cytosolic activity) was enriched by ion exchange chromatography after depletion of annexins by Ca2+-dependent phospholipid affinity chromatography. The fusion activity co-purified with a 10,14-kDa dimer identified as leukocyte L1 (which was non-fusogenic), along with an approximately 36-kDa protein. This protein was identified as glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by amino-terminal sequencing, and the fusion activity was verified using commercially available GAPDH. GAPDH may play an important role in degranulation because it is as potent as annexin I on a mass basis and may constitute up to 25% of the total cytosolic fusion activity of the neutrophil.