TARGETED DISRUPTION OF GUANOSINE DIPHOSPHATE-DISSOCIATION INHIBITOR FOR RHO-RELATED PROTEINS, GDID4 - NORMAL HEMATOPOIETIC DIFFERENTIATION BUT SUBTLE DEFECT IN SUPEROXIDE PRODUCTION BY MACROPHAGES DERIVED FROMIN-VITRO EMBRYONAL STEM-CELL DIFFERENTIATION

The Rho subfamily of small guanosine triphosphate (GTP)-binding protei ns, through their role in cytoskeletal organization, is involved in di verse cellular functions, including cell motility and morphologic chan ges during differentiation. pac also has a special role in the product ion of superoxide, a key component in phagocytic antimicrobial functio n, Guanosine diphosphate (GDP)-dissociation inhibitors (GDIs) belong t o one of three classes of proteins that regulate the critical cycling of GTP-binding proteins between the inactive and active states, Two ho mologous GDIs for the Rho subfamily have been identified. GDID4 is pre ferentially expressed in hematopoietic cells, while RhoGDI is ubiquito usly expressed, Whether different physiologic functions are subserved by the two GDIs is unknown, We have derived embryonal stem (ES) cells with targeted disruption of both alleles of the GDID4 gene and examine d hematopoiesis and phagocytic functions of macrophages derived from i n vitro ES-cell differentiation. GDID4(-/-) ES cells develop like wild type cells into colonies that contain heterogeneous populations of pro genitor cells and differentiated erythromyeloid cells. GDID4(-/-) cell s express no GDID4 protein, but have normal levels of RhoGDI, GDID4-/- macrophages phagocytose yeasts and antibody-opsonized erythrocytes as effectively as wild-type macrophages. However, a slight but consisten t reduction in their capacity to generate superoxide was observed, whi ch suggests new insight into the cellular role of GDID4, The minimal p henotypic effect of a loss of function of GDID4 also indicates a signi ficant redundancy of function between GDIDC and RhoGDI, Their function al repertoire may be better revealed by a disruption of both genes, Th e use of hematopoietic cells derived in vitro from genotypically alter ed ES cells avoids the difficulties inherent in generating knockout an imals and is a useful complementary approach for evaluating the gene f unction. (C) 1996 by The American Society of Hematology.