Differentiated blood cells arise via a series of commitment steps which progressively narrow the proliferative capacity and potential for cell diversity of the pluripotent hematopoietic stem cell. Evidence from study of both normal and leukemic cells suggests that a crucial step in neutrophil maturation occurs in the transition from the promyelocyte to the myelocyte stage. This transition is marked by a change in the granule content of the developing granulocyte. The promyelocyte stage is characterized by primary (""""""""nonspecific"""""""") granules which persist in decreasing numbers in the later phases of both neutrophil and monocyte maturation. The transition to the myelocyte stage, marked by a loss of proliferative capacity and restriction of the capacity for alternative maturation, is associated with the acquisition of secondary (""""""""specific"""""""") granules. These secondary granules are thus a unique marker of commitment to terminal neutrophil differentiation. Consequently, the expression of genes responsible for the formation of secondary granules and their contents should provide a useful marker for the processes controlling normal neutrophil differentiation. I have recently cloned the gene for one of the secondary granule proteins, transcobalamin I. This gene will be used as a probe for the factors controlling the stage-specific coordinate expression of granule protein genes during granulopoiesis. Effort will be concentrated on the characterization of expression of the gene for transcobalamin I in normal bone marrow cells in contrast with its expression in leukemic cells and in cells from patients with neutrophil specific granule deficiency. The major thrust of this research effort will be to isolate and characterize both """"""""cis"""""""" (enhancer, promoter, silencer) nucleotide sequence elements near the TCI gene, and the """"""""trans"""""""" acting protein factors that interact with those elements. The stages of neutrophil maturation characterized by secondary granule protein gene expression are associated with a simultaneous suppression of primary granule protein gene expression. Consequently, further analysis of the factors controlling expression of TCI will include comparison to the factors controlling expression of the gene for myeloperoxidase, a primary granule protein. These studies should offer important insight into the process of granulocyte differentiation, and the failure of this process during leukemogenesis.
