Chemotaxin-dependent translocation of immunoreactive ADP-ribosyltransferase-1 to the surface of human neutrophil polymorphs

mRNA from human polymorphonuclear neutrophil leucocytes (PMNs) was probed w ith cDNA encoding human skeletal muscle arginine-specific ADP-ribosyltransf erase (ART1). A single 2.6-kb transcript was identified, which was similar in size to that observed in human skeletal muscle RNA. An 872-bp cDNA fragm ent, corresponding to the amino acid sequence of the processed human skelet al muscle enzyme, was generated by reverse transcription-PCR amplification of RNA from human PMNs, and was found to be identical to the ART1 cDNA deri ved from human skeletal muscle. ART1 was expressed as a fusion protein with glutathione S-transferase (GST) in insect cells, and antibodies were raise d against the fusion protein in a rabbit. Following removal of GST immunore activity by immunoprecipitation, these antibodies were used to measure the abundance of immunoreactive ART1 on the surface of PMNs. Exposure of PMNs t o formyl-Met-Leu-Phr (FMLP) was followed by a rapid increase in the abundan ce of cell surface ART1 (T-1/2 = 1.9 min), and the concentration of FMLP fo r half-maximum response was 28.6 nM. Similar responses were observed after exposure of the cells to platelet-activating factor or interleukin-8, and w e conclude that some of the effects of these chemotaxins are mediated by tr anslocation of an intracellular pool of ART1 to its site of catalytic activ ity on the outer aspect of the plasma membrane.