Modulation of CD11b/CD18 Adhesive Activity by Its Extracellular, Membrane-Proximal Regions

Xiong, Ye; Chen, Jian; Zhang, Li

doi:10.4049/jimmunol.171.2.1042

Cited by 27 publications

(24 citation statements)

References 53 publications

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“…CHO-K1 cells stably transfected to express CD11b and CD18 (CHO CR3 ϩ ) or stably mock transfected with a neomycin resistance vector as control (CHO) were obtained as a kind gift from Douglas Golenbock and grown as described previously (31). K562 cells stably transfected to express CD11b and CD18 (K562 CR3 ϩ ) or K562 control cells (K562) were kind gifts of Li Zhang and were established and maintained as described previously (59). Surface expression of CD11b and CD18 was monitored by flow cytometry intermittently during the experiments to ensure stable coexpression using PE-conjugated MAb ICRF44 to CD11b and FITC-conjugated MAb 6.7 to CD18.…”

Section: Methodsmentioning

confidence: 99%

Role of CD11b/CD18 in the Process of Intoxication by the Adenylate Cyclase Toxin of Bordetella pertussis

et al. 2012

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The adenylate cyclase toxin (ACT) of Bordetella pertussis does not require a receptor to generate intracellular cyclic AMP (cAMP) in a broad range of cell types. To intoxicate cells, ACT binds to the cell surface, translocates its catalytic domain across the cell membrane, and converts intracellular ATP to cAMP. In cells that express the integrin CD11b/CD18 (CR3), ACT is more potent than in CR3-negative cells. We find, however, that the maximum levels of cAMP accumulation inside CR3-positive and -negative cells are comparable. To better understand how CR3 affects the generation of cAMP, we used Chinese hamster ovary and K562 cells transfected to express CR3 and examined the steps in intoxication in the presence and absence of the integrin. The binding of ACT to cells is greater in CR3-expressing cells at all concentrations of ACT, and translocation of the catalytic domain is enhanced by CR3 expression, with ϳ80% of ACT molecules translocating their catalytic domain in CR3-positive cells but only 25% in CR3-negative cells. Once in the cytosol, the unregulated catalytic domain converts ATP to cAMP, and at ACT concentrations >1,000 ng/ml, the intracellular ATP concentration is <5% of that in untreated cells, regardless of CR3 expression. This depletion of ATP prevents further production of cAMP, despite the CR3-mediated enhancement of binding and translocation. In addition to characterizing the effects of CR3 on the actions of ACT, these data show that ATP consumption is yet another concentration-dependent activity of ACT that must be considered when studying how ACT affects target cells.T he adenylate cyclase toxin (ACT) of Bordetella pertussis, the causative agent of whooping cough, is a single polypeptide composed of an N-terminal, 400-amino-acid adenylate cyclase (AC) enzymatic domain and a 1,306-amino-acid cell-binding domain homologous to the repeats-in-toxin (RTX) family of calcium-binding, pore-forming bacterial protein toxins (2,16,17,26,32). The AC domain converts ATP to cAMP in a high-turnover reaction that is stimulated by eukaryotic calmodulin (18). The RTX component forms oligomeric pores in cell membranes and serves as the cell binding domain (55, 58). To generate cyclic AMP (cAMP), ACT binds to cells, translocates the catalytic domain across the cytoplasmic membrane without endocytosis or macropinocytosis (11,19,25), and catalyzes the conversion of intracellular ATP to cAMP.Several cytotoxic effects for ACT have been identified, starting with the discovery that toxin-generated cAMP inhibits core functions of neutrophils and macrophages, such as the generation of an oxidative burst and killing of phagocytized bacteria (8, 49). ACT elicits additional cAMP-mediated effects in a variety of eukaryotic cells; these include apoptotic cell death, dysfunction of the cell cytoskeleton, cell cycle arrest, chloride secretion from polarized epithelial cells, and dysregulation of the adaptive immune system (8,11,22,30,34,47,49,50,54). To generate cAMP, the AC enzyme reaction consumes cellular ATP, further contrib...

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Section: Methodsmentioning

confidence: 99%

Role of CD11b/CD18 in the Process of Intoxication by the Adenylate Cyclase Toxin of Bordetella pertussis

et al. 2012

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“…CD11b/CD18-expressing HEK293 cells and mock-transfected HEK293 cells were generated previously (42). Rabbit polyclonal anti-CD18 cytoplasmic tail antibody ARC22 was prepared as described previously (43). Mannose-binding protein-invasin was provided by J.M.…”

Section: Antibodies and Reagents Mab F4/80 Was From Serotec; Mab M1/mentioning

confidence: 99%

“…Preparation of lipid rafts was accomplished as previously described (43). Murine macrophage-like RAW264 cells were grown to 90% confluence in 150 mm Petri-dishes.…”

Section: Antibodies and Reagents Mab F4/80 Was From Serotec; Mab M1/mentioning

confidence: 99%

The efficacy of activated protein C in murine endotoxemia is dependent on integrin CD11b

Cao¹,

Gao²,

Li³

et al. 2010

J. Clin. Invest.

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116

131

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Activated protein C (APC), the only FDA-approved biotherapeutic drug for sepsis, possesses anticoagulant, antiinflammatory, and barrier-protective activities. However, the mechanisms underlying its antiinflammatory functions are not well defined. Here, we report that the antiinflammatory activity of APC on macrophages is dependent on integrin CD11b/CD18, but not on endothelial protein C receptor (EPCR). We showed that CD11b/CD18 bound APC within specialized membrane microdomains/lipid rafts and facilitated APC cleavage and activation of protease-activated receptor-1 (PAR1), leading to enhanced production of sphingosine-1-phosphate (S1P) and suppression of the proinflammatory response of activated macrophages. Deletion of the γ-carboxyglutamic acid domain of APC, a region critical for its anticoagulant activity and EPCR-dependent barrier protection, had no effect on its antiinflammatory function. Genetic inactivation of CD11b, PAR1, or sphingosine kinase-1, but not EPCR, abolished the ability of APC to suppress the macrophage inflammatory response in vitro. Using an LPS-induced mouse model of lethal endotoxemia, we showed that APC administration reduced the mortality of wild-type mice, but not CD11b-deficient mice. These data establish what we believe to be a novel mechanism underlying the antiinflammatory activity of APC in the setting of endotoxemia and provide clear evidence that the antiinflammatory function of APC is distinct from its barrier-protective function and anticoagulant activities.

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“…CD18 expression on BMSSCs was achieved by viral infection (50-70% efficiency) using our published methods (17) and was verified by FACS analysis using mAb C71͞16 and by immunoblot using a polyclonal Ab against the cytoplasmic tail of CD18.…”

Section: Retroviral-mediated Cd18 Expression In Bmsscsmentioning

confidence: 99%

Defective osteogenesis of the stromal stem cells predisposes CD18-null mice to osteoporosis

Miura

Gronthos

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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Osteogenesis by the bone marrow stromal stem cells (BMSSCs) supports continuous bone formation and the homeostasis of the bone marrow microenvironment. The mechanism that controls the proliferation and differentiation of BMSSCs is not fully understood. Here, we report that CD18, a surface protein present primarily on hematopoietic cells, but not on differentiated mesenchymal cells, is expressed by the stromal stem cells and plays a critical role in the osteogenic process. Constitutive expression of CD18 on BMSSCs using a retroviral promoter significantly enhances bone formation in vivo, whereas genetic inactivation of CD18 in mice leads to defective osteogenesis due to decreased expression of the osteogenic master regulator Runx2͞Cbfa1. The defective osteogenesis of the CD18-null BMSSCs can be restored by expressing full-length, but not cytoplasmic domain-truncated, CD18. Radiographic analyses with dual-energy x-ray absorptiometry and 3D microcomputed tomography show that mice lacking CD18 have decreased bone mineral density and exhibit certain features of osteoporosis. Altogether, this work demonstrates that CD18 functions critically in the osteogenesis of BMSSCs, and thus lack of CD18 expression in the leukocyte adhesion deficiency patients may predispose them to osteoporosis.integrin ͉ leukocyte adhesion deficiency ͉ bone

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Modulation of CD11b/CD18 Adhesive Activity by Its Extracellular, Membrane-Proximal Regions

Cited by 27 publications

References 53 publications

Role of CD11b/CD18 in the Process of Intoxication by the Adenylate Cyclase Toxin of Bordetella pertussis

Role of CD11b/CD18 in the Process of Intoxication by the Adenylate Cyclase Toxin of Bordetella pertussis

The efficacy of activated protein C in murine endotoxemia is dependent on integrin CD11b

Defective osteogenesis of the stromal stem cells predisposes CD18-null mice to osteoporosis

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