We propose to characterize the G-protein genes which are expressed in human myeloid cells. These genes encode a family of proteins with a pervasive role in regulating the function of eucaryotic cells, including hematopoietic cells. Indirect evidence suggests that the G-proteins may be involved in the regulation of growth and differentiation of myeloid cells. Dr. Melvin I. Simon and his colleagues at Caltech have recently isolated cDNA clones for the alpha, beta, and gamma subunits of a bovine retinal G-protein, transducin. These cDNA clones, and oligonucleotide probes, will be used to study the pattern of G- protein gene expression in human myeloid cells. Initially, we will isolate cDNA clones corresponding to G-protein gene transcripts from a cDNA library generated from myeloid cell RNA. From the sequence of the cDNAs we will synthesize immunogenic peptides, then generate antibodies. Using nucleotide, antibody, and metabolic probes, we will identify G-protein genes which are expressed in myeloid cells and examine the role of these proteins in myeloid cell function. We will characterize the pattern of expression of the G-protein genes and their protein products in relation to cellular growth and differentiation of myeloid cells. By generation of antisense RNA, we will ablate the synthesis of G protein subunits in myeloid cells. We will then isolate genomic clones for those G-protein genes which are important to myeloid cell function. Increasing evidence suggests that somatic mutations occur in most leukemias (1,2). However, the mechanisms by which chromosomal translocations and alterations in oncogenes lead to abnormal growth remain obscure. We hope to gain some insight into the pathophysiology of abnormal growth and differentiation in myelogenous leukemia by examining the expression of G- protein genes in normal and leukemic myeloid cells. This proposal represent an ideal collaboration between a physician scientist and a basic researcher, as it fosters an interaction of complementary fields of expertise. Dr. Amatruda's previous studies on myeloid differentiation have prepared him to make full use of the excellent research environment which exists in Dr. Simon's laboratory and in the Caltech Division of Biology. In this setting, he will have a unique opportunity to learn the principles of molecular biology and apply them to the study of human disease.
