CR3 complement receptor: cloning and characterization in rainbow trout.
ABSTRACT The beta 2 integrin CR3 is a leukocyte adhesion heterodimeric glycoprotein which functions both as receptor for iC3b and in several cell-cell and cell-substrate adhesion interactions. In order to elucidate the molecular evolution of the CR3 receptor, here we report the cloning and characterization of its beta2 (CD18) and aM (CD11b) subunits in rainbow trout (Oncorhynchus mykiss). The predicted polypeptide sequences of trout CD18 and CD11b-like exhibit 50, 49, or 61% and 25, 25, or 30% identity with human, mouse, and zebrafish orthologs, respectively. The 'domain' architecture of trout CD18 and CD11b-like subunits retains several characteristics of the mammalian ortholog proteins, such as cysteine-rich regions, N-linked glycosylation sites and several proposed domains and signal sequences (von Willebrand factor type A, Integrin alpha, Integrin B tail, EGF, and Transmembrane domain). The tissue expression profiles of trout CR3 subunits diverge from those of mammalian counterparts, showing the kidney as the main source of the trout CD18 and CD11b-like mRNA transcripts. This is the first report of cloning and characterization of the CR3 receptor in low vertebrates.
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Article: Integrin regulation.[show abstract] [hide abstract]
ABSTRACT: Integrin signaling is bidirectional. 'Inside-out' signals regulate integrin affinity for adhesive ligands, and ligand-dependent 'outside-in' signals regulate cellular responses to adhesion. Integrin extracellular domains are yielding to high-resolution structural analyses, and intracellular proteins involved in integrin signaling are being identified. However, a key unresolved question is how integrins propagate signals across the plasma membrane.Current Opinion in Cell Biology 11/2005; 17(5):509-16. · 11.41 Impact Factor
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ABSTRACT: Integrin-type complement receptors play pivotal roles in the effector mechanisms of the complement system. Previously, we identified an integrin alpha subunit, alpha(Hr1), from the solitary ascidian, Halocynthia roretzi, which is involved in the complement-dependent phagocytic activities of ascidian hemocytes. To identify integrin beta subunits that pair with alpha(Hr1) to compose ascidian complement receptors, genes encoding beta subunits were cloned and characterized for their binding property to alpha(Hr1). Using degenerate primers and RT-PCR, two integrin beta transcripts (beta(Hr1) and beta(Hr2)) were isolated from H. roretzi hemocyte total RNA and the entire coding sequences of both cDNA species were determined. The putative primary structure of each ascidian gene product retained domains characteristic for integrin beta subunits. Phylogenetic analysis revealed that beta(Hr1) and beta(Hr2) are located outside of vertebrate integrin beta groups, comprising an independent cluster specific for the ascidian lineage. The alpha(Hr1), beta(Hr1) and beta(Hr2) subunits all showed hemocyte-specific expression on Northern blot analysis, and recombinant proteins of both beta subunits could bind to alpha(Hr1) on insect cells. The beta(Hr1) subunit was expressed especially on the surface of ascidian phagocytic hemocytes, such as phago-amoebocytes. In the immunoprecipitation analysis of ascidian hemocytes using anti-beta(Hr1) antiserum, alpha(Hr1) was coprecipitated with beta(Hr1). These observations showed that beta(Hr1), and possibly beta(Hr2) too, binds to alpha(Hr1) to comprise integrin molecules on ascidian hemocytes, which act as ancestral forms of complement receptors in the primitive complement system of ascidians.Immunogenetics 04/2004; 55(12):836-44. · 2.89 Impact Factor
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ABSTRACT: Despite the identification and characterization of several distinct ligands for the leukocyte integrin (CD11/CD18) family of adhesion receptors, little is known about the structural regions on these molecules that mediate ligand recognition. In this report, we use alpha subunit chimeras of Mac-1 (CD11b/CD18) and p150,95 (CD11c/CD18), and an extended panel of newly generated and previously characterized mAbs specific to the alpha chain of Mac-1 to map the binding sites for four distinct ligands for Mac-1: iC3b, fibrinogen, ICAM-1, and the as-yet uncharacterized counter-receptor responsible for neutrophil homotypic adhesion. Epitopes of mAbs that blocked ligand binding were mapped with the chimeras and used to localize the ligand recognition sites because the data obtained from functional assays with the Mac-1/p150,95 chimeras were not easily interpreted. Results show that the I domain on the alpha chain of Mac-1 is an important recognition site for all four ligands, and that the NH2-terminal and perhaps divalent cation binding regions but not the COOH-terminal segment may contribute. The recognition sites in the I domain appear overlapping but not identical as individual Mac-1-ligand interactions are distinguished by the discrete patterns of inhibitory mAbs. Additionally, we find that the alpha subunit NH2-terminal region and divalent cation binding region, despite being separated by over 200 amino acids of the I domain, appear structurally apposed because three mAbs require the presence of both of these regions for antigenic reactivity, and chimeras that contain the NH2 terminus of p150,95 require the divalent cation binding region of p150,95 to associate firmly with the beta subunit.The Journal of Cell Biology 03/1993; 120(4):1031-43. · 10.82 Impact Factor