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Costimulation of T cell activation by integrin-associated protein (CD47) is an adhesion-dependent, CD28-independent signaling pathway.

Reinhold MI, Lindberg FP, Kersh GJ, Allen PM, Brown EJ - J. Exp. Med. (1997)

Bottom Line: Here, we show that mAbs recognizing IAP can enhance proliferation of primary human T cells in the presence of low levels of anti-CD3, but have no effect on T cell proliferation on their own.IAP costimulation does not require CD28.We hypothesize that anti-IAP generates the costimulatory signal because it modulates interactions of the IgV domain with other plasma membrane molecules; this in turn activates effector functions of the multiply membrane-spanning domain of IAP.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
The integrin-associated protein (IAP, CD47) is a 50-kD plasma membrane protein with a single extracellular immunoglobulin variable (IgV)-like domain, a multiply membrane-spanning segment, and alternatively spliced short cytoplasmic tails. On neutrophils, IAP has been shown to function in a signaling complex with beta 3 integrins. However, the function of IAP on T cells, which express little or no beta 3 integrin, is not yet defined. Here, we show that mAbs recognizing IAP can enhance proliferation of primary human T cells in the presence of low levels of anti-CD3, but have no effect on T cell proliferation on their own. Together with suboptimal concentrations of anti-CD3, engagement of IAP also enhances IL-2 production in Jurkat cells, an apparently integrin-independent function of IAP. Nonetheless, costimulation by IAP ligation requires cell adhesion. IAP costimulation does not require CD28. Furthermore, anti-IAP, but not anti-CD28, synergizes with suboptimal anti-CD3 to enhance tyrosine phosphorylation of the CD3 zeta chain and the T cell-specific tyrosine kinase Zap70. Ligation of human IAP transfected into the hemoglobin-specific 3.L2 murine T cell hybridoma costimulates activation for IL-2 secretion both with anti-CD3 and with antigenic peptides on antigen-presenting cells (APCs). Moreover, ligation of IAP but not CD28 can convert antagonist peptides into agonists in 3.L2 cells. Using costimulation by IAP ligation as an assay to analyze the structure-function relationships in IAP signaling, we find that both the extracellular and multiply membrane-spanning domains of IAP are necessary for synergy with the antigen receptor, but the alternatively spliced cytoplasmic tails are not. These data demonstrate that IAP ligation initiates an adhesion-dependent costimulatory pathway distinct from CD28. We hypothesize that anti-IAP generates the costimulatory signal because it modulates interactions of the IgV domain with other plasma membrane molecules; this in turn activates effector functions of the multiply membrane-spanning domain of IAP. This model may have general significance for how IAP functions in cell activation.

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hIAP and chimera expression in Jurkat and 3.L2 subclones.  Expression of native hIAP and chimeric constructs was determined by  staining with mouse anti–hIAP IgG1 (2E11) or rat anti–murine CD8α  (dotted lines), or isotype-matched control (7G2 or 313, respectively, solid  lines) mAbs as described in Materials and Methods. Shown are profiles of  one of the transfected Jurkat or 3.L2 clones, with similar levels of expression in the second clone transfected.
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Figure 4: hIAP and chimera expression in Jurkat and 3.L2 subclones. Expression of native hIAP and chimeric constructs was determined by staining with mouse anti–hIAP IgG1 (2E11) or rat anti–murine CD8α (dotted lines), or isotype-matched control (7G2 or 313, respectively, solid lines) mAbs as described in Materials and Methods. Shown are profiles of one of the transfected Jurkat or 3.L2 clones, with similar levels of expression in the second clone transfected.

Mentions: To begin to understand IAP function in T cell costimulation, we transfected form 2 of human IAP (hIAP), which is the predominant IAP form found in leukocytes, into two independent clones of the hemoglobin-specific murine T cell hybridoma 3.L2 (Fig. 4) (17). Initial experiments using the 3.L2 clones showed that mAbs recognizing CD28 costimulated IL-2 production with suboptimal antiCD3, indicating that this murine hybridoma was responsive to costimulatory signals. Coligation of hIAP with antihIAP mAbs, which do not cross-react with mouse IAP, and suboptimal anti-CD3 also resulted in enhanced IL-2 production over control mAb (Fig. 5 A).


Costimulation of T cell activation by integrin-associated protein (CD47) is an adhesion-dependent, CD28-independent signaling pathway.

Reinhold MI, Lindberg FP, Kersh GJ, Allen PM, Brown EJ - J. Exp. Med. (1997)

hIAP and chimera expression in Jurkat and 3.L2 subclones.  Expression of native hIAP and chimeric constructs was determined by  staining with mouse anti–hIAP IgG1 (2E11) or rat anti–murine CD8α  (dotted lines), or isotype-matched control (7G2 or 313, respectively, solid  lines) mAbs as described in Materials and Methods. Shown are profiles of  one of the transfected Jurkat or 3.L2 clones, with similar levels of expression in the second clone transfected.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2211576&req=5

Figure 4: hIAP and chimera expression in Jurkat and 3.L2 subclones. Expression of native hIAP and chimeric constructs was determined by staining with mouse anti–hIAP IgG1 (2E11) or rat anti–murine CD8α (dotted lines), or isotype-matched control (7G2 or 313, respectively, solid lines) mAbs as described in Materials and Methods. Shown are profiles of one of the transfected Jurkat or 3.L2 clones, with similar levels of expression in the second clone transfected.
Mentions: To begin to understand IAP function in T cell costimulation, we transfected form 2 of human IAP (hIAP), which is the predominant IAP form found in leukocytes, into two independent clones of the hemoglobin-specific murine T cell hybridoma 3.L2 (Fig. 4) (17). Initial experiments using the 3.L2 clones showed that mAbs recognizing CD28 costimulated IL-2 production with suboptimal antiCD3, indicating that this murine hybridoma was responsive to costimulatory signals. Coligation of hIAP with antihIAP mAbs, which do not cross-react with mouse IAP, and suboptimal anti-CD3 also resulted in enhanced IL-2 production over control mAb (Fig. 5 A).

Bottom Line: Here, we show that mAbs recognizing IAP can enhance proliferation of primary human T cells in the presence of low levels of anti-CD3, but have no effect on T cell proliferation on their own.IAP costimulation does not require CD28.We hypothesize that anti-IAP generates the costimulatory signal because it modulates interactions of the IgV domain with other plasma membrane molecules; this in turn activates effector functions of the multiply membrane-spanning domain of IAP.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
The integrin-associated protein (IAP, CD47) is a 50-kD plasma membrane protein with a single extracellular immunoglobulin variable (IgV)-like domain, a multiply membrane-spanning segment, and alternatively spliced short cytoplasmic tails. On neutrophils, IAP has been shown to function in a signaling complex with beta 3 integrins. However, the function of IAP on T cells, which express little or no beta 3 integrin, is not yet defined. Here, we show that mAbs recognizing IAP can enhance proliferation of primary human T cells in the presence of low levels of anti-CD3, but have no effect on T cell proliferation on their own. Together with suboptimal concentrations of anti-CD3, engagement of IAP also enhances IL-2 production in Jurkat cells, an apparently integrin-independent function of IAP. Nonetheless, costimulation by IAP ligation requires cell adhesion. IAP costimulation does not require CD28. Furthermore, anti-IAP, but not anti-CD28, synergizes with suboptimal anti-CD3 to enhance tyrosine phosphorylation of the CD3 zeta chain and the T cell-specific tyrosine kinase Zap70. Ligation of human IAP transfected into the hemoglobin-specific 3.L2 murine T cell hybridoma costimulates activation for IL-2 secretion both with anti-CD3 and with antigenic peptides on antigen-presenting cells (APCs). Moreover, ligation of IAP but not CD28 can convert antagonist peptides into agonists in 3.L2 cells. Using costimulation by IAP ligation as an assay to analyze the structure-function relationships in IAP signaling, we find that both the extracellular and multiply membrane-spanning domains of IAP are necessary for synergy with the antigen receptor, but the alternatively spliced cytoplasmic tails are not. These data demonstrate that IAP ligation initiates an adhesion-dependent costimulatory pathway distinct from CD28. We hypothesize that anti-IAP generates the costimulatory signal because it modulates interactions of the IgV domain with other plasma membrane molecules; this in turn activates effector functions of the multiply membrane-spanning domain of IAP. This model may have general significance for how IAP functions in cell activation.

Show MeSH
Related in: MedlinePlus