Unanue. be labeled with anti-BM8, another macrophage subset-specific marker with hitherto undefined molecular antigenic structure. Recombinant expression of a F4/80 cDNA in heterologous cells confirmed this observation, indicating that the targets recognized by the F4/80 and BM8 monoclonal antibodies are identical. Macrophages and their precursors, blood monocytes, represent widely distributed populations of myeloid cells. Resident tissue macrophages display extensive functional and phenotypic heterogeneity shaped in large part by the tissue microenvironment, as do monocytes recruited to local sites in response to inflammatory and immune stimuli (16, 26). Immunohistochemical analyses of normal tissue sections and various tissues from infected or immunologically challenged mice have utilized panels of monoclonal antibodies which recognize phenotypic markers expressed by macrophage subpopulations, such as macrosialin (the mouse ortholog of CD68), sialoadhesin (mouse CD169), scavenger receptor (SRA-I and SRA-II), complement receptor type 3 (CR3, CD11b/CD18), and the F4/80 Alosetron glycoprotein (3, 5, 10, 16, 26, 35, 39, 43). Perhaps the most extensively utilized reagent for detection of mature tissue macrophages is the monoclonal antibody (MAb) F4/80, which recognizes the F4/80 molecule expressed at high levels on the surface of a range of cells including Kupffer cells in the liver, splenic red pulp macrophages, brain microglia, gut lamina propria, and Langerhans cells in the skin, as well as resident macrophages found throughout connective tissue and many other organs including the heart, kidney, and the reproductive and neuroendocrine systems (3, 21-23, 31, 36). The isolation of cDNA clones encoding the mouse F4/80 glycoprotein resulted in the molecular characterization of its unusual structure and led to the further identification of related leukocyte cell surface proteins (4, 6, 27, 31, 44, 45). The F4/80 molecule contains seven-transmembrane (TM7) regions, which anchor the protein in the cell membrane, and thereby shows similarity in this region to G-protein-coupled receptors. The F4/80 molecule shares overall structural homology to other members of the epidermal growth factor (EGF)-TM7 family, which includes EMR1 (the human ortholog of F4/80), CD97, EMR2, EMR3, and FIRE, which are differentially expressed on populations of leukocytes, as well as ETL, which is expressed by cardiomyocytes (4, 6, 19, 32, 44, 45). Alosetron The EGF-TM7 gene family cluster has been mapped to human chromosome 19p and syntenic regions within the mouse genome (7, 27, 44). These EGF-TM7 molecules are a subset of a larger group of TM7 molecules termed LNB-TM7, which are defined by amino acid sequence similarity within the TM7 region to members of G-protein-coupled receptor family B, yet have large N-terminal extracellular regions often containing a complex array of protein modules (44). The presence of such large and composite structures also suggests specialized functions for these molecules, possibly in addition to peptide receptor activities. To date, the function of F4/80 has not been determined, although speculation has suggested its involvement in macrophage adhesion events, either to matrix molecules FMN2 or Alosetron a ligand expressed on other cell types, a role in cell migration or as a G protein-coupled signaling component of macrophages (30, 44). Indeed, one member of the EGF-TM7 family, CD97, has been shown to bind CD55 (decay accelerating factor), a glycosylphosphatidylinositol-linked cell-surface receptor previously shown to regulate complement activation by catalyzing the dissociation of components of the C3 convertases (30, 44). This interaction was specific for a particular isoform of CD97 containing three EGF domains (EGF1, EGF2, and EGF5), suggesting that EGF-TM7 gene-splicing mechanisms regulate receptor-ligand pairing (20). Alosetron In fact, this represented the first demonstration of a TM7 molecule binding a cellular ligand, although the functional significance of this interaction has yet to be established. In the absence of information relating to the function of F4/80 and its ligand(s), we sought to address the functional consequences of deleting the gene in a mouse germ line. In parallel, we aimed to generate a mouse line that specifically expresses Cre recombinase (38) within F4/80+ macrophage populations, thereby allowing cell-specific targeted deletion of candidate locus. A.
