TIMING TARGETING AND SORTING OF AZUROPHIL GRANULE PROTEINS IN HUMAN MYELOID CELLS

Biosynthesis of 3 human granule proteins, myeloperoxidase, defensin an d lysozyme, all present in azurophil granules, was investigated in nor mal bone marrow cells and in the promyelocytic cell line HL-60 to see whether differences in timing of biosynthesis could explain the well e stablished differences in their subcellular localization in the mature neutrophil (targeting), and whether differences exist in the efficien cies by which granule proteins are retained in cells (sorting). Normal human bone marrow cells were separated into three bands by density gr adient centrifugation. Band 1 contains band and segmented cells, band 2 mainly myelocytes, metamyelocytes and some band cells, and band 3 my eloblasts and promyelocytes in addition to megakaryocytes and proeryth roblasts. Cells from these bands, as well as undifferentiated HL-60 ce lls, were pulsed with radiolabeled cysteine and methionine, and biosyn thesis of granule proteins was subsequently evaluated by immunoprecipi tation and quantified by phosphorimaging. Myeloperoxidase synthesis wa s maximal in cells from band 3 while defensin biosynthesis was maximal in cells from band 2. Lysozyme was synthesized in cells from all band s but was maximal in cells from band 2. These results are in agreement with our hypothesis that timing of biosynthesis determines the locali zation of individual granule proteins. While myeloperoxidase and defen sins were efficiently retained in immature cells (band 3), a significa nt fraction of lysozyme was routed out of the cells, showing that diff erences exist in the sorting of granule proteins between constitutive and regulated secretion. In addition, defensin was less efficiently re tained in cells from band 2 than from band 3, indicating that sorting mechanisms may depend on the stage of cell maturation.