title: Targeted disruption of guanosine diphosphate-dissociation inhibitor for Rho-related proteins, GDID4: normal hematopoietic differentiation but subtle defect in superoxide production by macrophages derived from in vitro embryonal stem cell differentiation.
abstract: The Rho subfamily of small guanosine triphosphate (GTP)-binding proteins, through their role in cytoskeletal organization, is involved in diverse cellular functions, including cell motility and morphologic changes during differentiation. Rac also has a special role in the production of superoxide, a key component in phagocytic antimicrobial function. Guanosine diphosphate (GDP)-dissociation inhibitors (GDIs) belong to one of three classes of proteins that regulate the critical cycling of GTP-binding proteins between the inactive and active states. Two homologous GDIs for the Rho subfamily have been identified. GDID4 is preferentially expressed in hematopoietic cells, while RhoGDI is ubiquitously 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 examined hematopoiesis and phagocytic functions of macrophages derived from in vitro ES-cell differentiation. GDID4-/- ES cells develop like wild-type cells into colonies that contain heterogeneous populations of progenitor cells and differentiated erythromyeloid cells. GDID4-/- cells 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 consistent reduction in their capacity to generate superoxide was observed, which suggests new insight into the cellular role of GDID4. The minimal phenotypic effect of a loss of function of GDID4 also indicates a significant redundancy of function between GDID4 and RhoGDI. Their functional repertoire may be better revealed by a disruption of both genes. The use of hematopoietic cells derived in vitro from genotypically altered ES cells avoids the difficulties inherent in generating knockout animals and is a useful complementary approach for evaluating the gene function.
authors.affiliation: Division of Hematology/Oncology, Beth Israel Hospital, Harvard Medical School, Boston, MA, USA.
authors.forename: J C
authors.initials: JC
authors.lastname: Guillemot
authors.forename: B A
authors.initials: BA
authors.lastname: Kruskal
authors.forename: C N
authors.initials: CN
authors.lastname: Adra
authors.forename: S
authors.initials: S
authors.lastname: Zhu
authors.forename: J L
authors.initials: JL
authors.lastname: Ko
authors.forename: P
authors.initials: P
authors.lastname: Burch
authors.forename: K
authors.initials: K
authors.lastname: Nocka
authors.forename: K
authors.initials: K
authors.lastname: Seetoo
authors.forename: E
authors.initials: E
authors.lastname: Simons
authors.forename: B
authors.initials: B
authors.lastname: Lim
mesh_terms: D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D004622:Embryo, Mammalian / Q000166:cytology; D005347:Fibroblasts / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism / Q000502:physiology; D018390:Gene Targeting; D020730:Guanine Nucleotide Dissociation Inhibitors; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006636:Histamine Release / Q000187:drug effects; D007073:Immunoglobulin E / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008264:Macrophages / Q000378:metabolism*; D008407:Mast Cells / Q000166:cytology; D051379:Mice; D015778:Minor Histocompatibility Antigens; D010587:Phagocytosis; D011506:Proteins / Q000235:genetics* / Q000502:physiology; D016897:Respiratory Burst; D019089:Stem Cell Factor / Q000494:pharmacology; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism*; D062949:rho Guanine Nucleotide Dissociation Inhibitor beta; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors
chemical_list: C569659:Arhgdib protein, mouse; D020730:Guanine Nucleotide Dissociation Inhibitors; D007476:Ionophores; D015778:Minor Histocompatibility Antigens; D011506:Proteins; D019089:Stem Cell Factor; D062949:rho Guanine Nucleotide Dissociation Inhibitor beta; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors; D013481:Superoxides; D007073:Immunoglobulin E; D000001:Calcimycin; D019204:GTP-Binding Proteins; D002118:Calcium