Integrin Binding to the Trimeric Interface of CD40L Plays a Critical Role in CD40/CD40L Signaling.

CD40L plays a major role in immune response and is a major therapeutic target for inflammation. Integrin α5ß1 and CD40 simultaneously bind to CD40L. It is unclear if α5ß1 and CD40 work together in CD40/CD40L signaling or how α5ß1 binds to CD40L. In this article, we describe that the integrin-binding site of human CD40L is predicted to be located in the trimeric interface by docking simulation. Mutations in the predicted integrin-binding site markedly reduced the binding of α5ß1 to CD40L. Several CD40L mutants defective in integrin binding were defective in NF-κB activation and B cell activation and suppressed CD40L signaling induced by wild-type CD40L; however, they still bound to CD40. These findings suggest that integrin α5ß1 binds to monomeric CD40L through the binding site in the trimeric interface of CD40L, and this plays a critical role in CD40/CD40L signaling. Integrin αvß3, a widely distributed vascular integrin, bound to CD40L in a KGD-independent manner, suggesting that αvß3 is a new CD40L receptor. Several missense mutations in CD40L that induce immunodeficiency with hyper-IgM syndrome type 1 (HIGM1) are clustered in the integrin-binding site of the trimeric interface. These HIGM1 CD40L mutants were defective in binding to α5ß1 and αvß3 (but not to CD40), suggesting that the defect in integrin binding may be a causal factor of HIGM1. These findings suggest that α5ß1 and αvß3 bind to the overlapping binding site in the trimeric interface of monomeric CD40L and generate integrin-CD40L-CD40 ternary complex. CD40L mutants defective in integrins have potential as antagonists of CD40/CD40L signaling.

A novel CD40LG deletion causes the hyper-IgM syndrome with normal CD40L expression in a 6-month-old child.

The X-linked hyper-IgM syndrome (XHIGM) is the most common form of HIGM. Patients are clinically diagnosed on the basis of recurrent sinopulmonary infections, accompanied with low levels of IgG and IgA, normal to elevated levels of IgM, and the presence of peripheral B cells. Here, we have reported a novel deletion of four nucleotides in CD40LG exon 3, c.375_378delCAAA, which led to a frameshift mutation with a premature stop codon, p.Asn101*126. The deletion resulted in a truncated protein, in which majority of the extracellular domain was lost. However, detection of surface CD40L was still possible as the intracellular, transmembrane, and part of the extracellular domains were not affected. This indicated that this mutation did not affect protein stability and that immunodetection of CD40L expression is not enough for the diagnosis of XHIGM. Our study strongly suggests that genetic diagnosis for XHIGM should always be performed when clinical data support this diagnosis and CD40L protein is present.

The role of CD40/CD40 ligand interactions in bone marrow granulopoiesis.

The CD40 ligand (CD40L) and CD40 are two molecules belonging to the TNF/TNF receptor superfamily, and their role in adaptive immune system has widely been explored. However, the wide range of expression of these molecules on hematopoietic as well as nonhematopoietic cells has revealed multiple functions of the CD40/CD40L interactions on different cell types and processes such as granulopoiesis. CD40 triggering on stromal cells has been documented to enhance the expression of granulopoiesis growth factors such as granulocyte-colony-stimulating factor (G-CSF) and granulocyte/monocyte-colony-stimulating factor (GM-CSF), and upon disruption of the CD40/CD40L-signaling pathway, as in the case of X-linked hyperimmunoglobulin M (IgM) syndrome (XHIGM), it can lead to neutropenia. In chronic idiopathic neutropenia (CIN) of adults, however, under the influence of an inflammatory microenvironment, CD40L plays a role in granulocytic progenitor cell depletion, providing thus a pathogenetic cause of CIN.

Impaired maintenance of naturally acquired T-cell memory to the meningococcus in patients with B-cell immunodeficiency.

The importance of T cells in the generation of antigen-specific B-cell immunity has been extensively described, but the role B cells play in shaping T-cell memory is uncertain. In healthy controls, exposure to Neisseria meningitidis in the upper respiratory tract is associated with the generation of memory T cells in the mucosal and systemic compartments. However, we demonstrate that in B cell-deficient subjects with X-linked agammaglobulinemia (XLA), naturally acquired T-cell memory responses to meningococcal antigens are reduced compared with healthy control patients. This difference is not found in T-cell memory to an obligate respiratory pathogen, influenza virus. Accordingly, we show that meningococcal antigens up-regulate major histocompatibility complex (MHC) class II, CD40, CD86/80 expression on mucosal and systemic associated B cells and that antigen presentation stimulates T-cell proliferation. A similar reduction in N meningitidis but not influenza antigen-specific T-cell memory was observed in subjects with X-linked hyper IgM syndrome (X-HIM), implicating the interaction of CD40-CD40L in this process. Together, these data implicate B cells in the induction and maintenance of T-cell memory to mucosal colonizing bacteria such as N meningitidis and highlight the importance of B cells beyond antibody production but as a target for immune reconstitution.

Poorly differentiated gastroenteropancreatic neuroendocrine carcinoma associated with X-linked hyperimmunoglobulin M syndrome.

Gastroenteropancreatic neuroendocrine tumors are uncommon tumors representing 2% of all gastrointestinal tumors. We report a case of a 21-year-old man with X-linked hyperimmunoglobulin M (hyper-IgM) syndrome who presented with diarrhea and jaundice. An ultrasound and magnetic resonance imaging showed multiple variable-sized lesions in the liver and peripancreatic lymphadenopathy. The morphologic and immunohistochemical features of the biopsies from the liver and lymph node were consistent with poorly differentiated neuroendocrine carcinoma. Hyper-IgM syndrome is a rare primary immunodeficiency disease characterized by low serum IgG, IgA, and IgE levels with normal or elevated IgM levels. These patients are at a higher risk for developing malignancies, particularly adenocarcinoma of the gastrointestinal tract and lymphoma. A review of the literature of gastroenteropancreatic neuroendocrine tumors is presented with the discussion of a possible relationship of these tumors with immunodeficiency.

c-Rel plays a key role in deficient activation of B cells from a non-X-linked hyper-IgM patient.

Our previous results demonstrated that B cells from a patient (pt1) with non-X-linked hyper-IgM syndrome (HIGM) possess an atypical CD23(lo) phenotype that is unaffected by CD40-mediated activation. To investigate the molecular mechanism underlying defective CD23 expression in pt1 B cells, we used lymphoblastoid cell lines that express LMP1 under the control of a tetracycline-inducible promoter (LCL(tet)). Our analysis revealed that the CD23(lo) phenotype in the pt1-LCL(tet) cells is a direct consequence of diminished CD23 transcription. We demonstrate a marked decrease in c-Rel-containing complexes that bind to the proximal CD23a/b promoters in pt1-LCL(tet) extracts, resulting from an overall lower expression of c-Rel in pt1-LCL(tet) cells. Analysis of c-Rel mRNA revealed relatively equal amounts in pt1-LCL(tet) and control LCL(tet) cells, indicating that diminished c-Rel protein expression is unrelated to decreased transcription. Finally, a critical role for c-Rel in CD23 regulation was demonstrated by effectively altering c-Rel expression that resulted in the direct modulation of CD23 surface expression. Collectively, these findings demonstrate that low levels of c-Rel are the underlying cause of aberrant CD23 expression in pt1 B cells and are likely to play a critical role in the pathophysiology of this form of HIGM.