Roland Geisberger

The riddle of the dual expression of IgM and IgD.

Signalling through the B cell antigen receptor (BCR) is required for peripheral B lymphocyte maturation, maintenance, activation and silencing. In mature B cells, the antigen receptor normally consists of two isotypes, membrane IgM and IgD (mIgM, mIgD). Although the signals initiated from both isotypes differ in kinetics and intensity, in vivo, the BCR of either isotype seems to be able to compensate for the loss of the other, reflected by the mild phenotypes of mice deficient for mIgM or mIgD. Thus, it is still unclear why mature B cells need expression of mIgD in addition to mIgM. In the current review we suggest that the view that IgD has a simpIy definable function centred around the basic signalling function should be replaced by the assumption that IgD fine tunes humoral responses, modulates B cell selection and homeostasis and thus shapes the B cell repertoire, defining IgD to be a key modulator of the humoral immune response.

T cell exhaustion: from pathophysiological basics to tumor immunotherapy

The immune system is capable of distinguishing between danger- and non-danger signals, thus inducing either an appropriate immune response against pathogens and cancer or inducing self-tolerance to avoid autoimmunity and immunopathology. One of the mechanisms that have evolved to prevent destruction by the immune system, is to functionally silence effector T cells, termed T cell exhaustion, which is also exploited by viruses and cancers for immune escape In this review, we discuss some of the phenotypic markers associated with T cell exhaustion and we summarize current strategies to reinvigorate exhausted T cells by blocking these surface marker using monoclonal antibodies.

Somatic diversity of the immunoglobulin repertoire is controlled in an isotype‐specific manner

We have studied two aspects of the IgE immune response. First, we have compared the kinetics of the IgE response to the T cell‐dependent antigen ph‐Ox coupled to ovalbumin with that of the IgG1 response and we have assessed the quality of the IgE response. Second, we have studied the generation of somatic diversity, understood as the combined effect of somatic mutation and the selection of D(iversity) and J(oining) elements, in germinal center B cells at the molecular level, using the germ‐line sequence of the prototype anti‐ph‐Ox heavy chain variable element VHOx1 as reference. We evaluated sequences derived from μ‐, γ1‐ and ϵ‐variable elements and showed that somatic diversification was different for all isotypes studied. We further compared the IgE responses of wild‐type mice with those of mice expressing a truncated cytoplasmic IgE tail (IgEKVKΔtail). IgEKVKΔtail mice showed a more diverse sequence pattern. We corroborated previous results suggesting that short CDR3 regions are indicative for high‐affinity antibodies by measuring relative affinities of phage‐expressed Fab fragments with prototype long and short CDR3 regions. Therefore, the composition of the antigen‐receptor is responsible for the selection process and the expansion of antigen‐specific cells, leading to an isotype‐specific antibody repertoire.

Models of signal transduction through the B-cell antigen receptor

From the numerous surface markers of a B lymphocyte, the B-cell antigen receptor (BCR) complex is probably the most powerful marker influencing the developmental processes of the cell. The BCR consists of the membrane-bound immunoglobulin (mIg) but, depending on the state of differentiation, may be associated with a couple of other transmembrane proteins, most notably Igα (CD79a) and Igβ (CD79b).1 The N-terminal end of the mIg harbours the antigen-binding site, characterized by an incredibly high potential for diversification and built up by the variable regions of heavy and light chains. Followed by three or four constant domains, depending on the selected immunoglobulin-isotype, the mIg finally expresses two further domains: a transmembrane domain and a cytoplasmic tail, both of which vary in their isotype-specific amino acid composition.2 So far, the sheath proteins Igα and Igβ are known as the signal transduction component of the BCR complex, connecting the antigen receptor to the tyrosine phosphorylation pathways in the cell. All isotypes of mIg can form a complex with Igα and Igβ,3 indicating an involvement of all isotypes in the signal transduction pathway. Venkitaraman et al.3 showed that for mIgG and mIgD the Igα/Igβ sheath is not required for surface expression. However, Igα/Igβ is the minimum requirement for signal transduction4 and, in the case of IgM, is responsible for the release from intracellular retention sites.5 Igα is expressed by the mb-1 gene and is a 32 000 MW glycoprotein. Igβ (B29 gene) can be expressed in two different isoforms of 37 000 and 39 000 MW, respectively. Interestingly, Igα can be differentially glycosylated. Pogue and Goodnow6 suggested that the extracellular spacer domain of mIgD is necessary and sufficient to confer the mIgD-specific glycosylation pattern of the mb-1 gene.6 However, it remains to be elucidated if and how alternative glycosylation of mb-1 may affect signalling competence or internalization.

Chemotherapy-induced augmentation of T cells expressing inhibitory receptors is reversed by treatment with lenalidomide in chronic lymphocytic leukemia

While T-cell dysfunction occurring alongside chronic lymphocytic leukemia (CLL) is well documented (reviewed by Pleyer et al. [1][1] and Hamblin et al. [2][2]), it was recently reported that also T-cell exhaustion is associated with CLL, reflected in the presence of PD-1+ CLL T cells and higher

HS1-Associated Protein X-1 Interacts with Membrane-Bound IgE: Impact on Receptor-Mediated Internalization

Engagement of the BCR triggers signals that control affinity maturation, memory induction, differentiation, and various other physiological processes in B cells. In previous work, we showed that truncation of the cytoplasmic tail of membrane-bound Ig (mIg)E in vivo resulted in lower serum IgE levels, decreased numbers of IgE-secreting plasma cells, and the abrogation of specific secondary responses correlating with a defect in the selection of high-affinity Abs during the germinal center reaction. We concluded that the Ag receptor is necessary at all times during Ab responses not only for the maturation process, but also for the expansion of Ag-specific B cells. Based on these results, we asked whether the cytoplasmic tail of mIgE, or specific proteins binding the cytoplasmic tail in vivo commit a signal transduction accompanying the B cell along its differentiation process. In this study, we present the identification of HS1-associated protein X-1 as a novel protein interacting with the cytoplasmic tail of mIgE. ELISA, surface plasmon resonance analysis, and coimmunoprecipitation experiments confirmed the specific interaction in vitro. In functional assays, we clearly showed that HS1-associated protein X-1 expression levels influence the efficiency of BCR-mediated Ag internalization.

Fludarabine modulates composition and function of the T cell pool in patients with chronic lymphocytic leukaemia

The combination of cytotoxic treatment with strategies for immune activation represents an attractive strategy for tumour therapy. Following reduction of high tumour burden by effective cytotoxic agents, two major immune-stimulating approaches are being pursued. First, innate immunity can be activated by monoclonal antibodies triggering antibody-dependent cellular cytotoxicity. Second, tumour-specific T cell responses can be generated by immunization of patients with peptides derived from tumour antigens and infused in soluble form or loaded onto dendritic cells. The choice of cytotoxic agents for such combinatory regimens is crucial since most substances such as fludarabine are considered immunosuppressive while others such as cyclophosphamide can have immunostimulatory activity. We tested in this study whether fludarabine and/or cyclophosphamide, which represent a very effective treatment regimen for chronic lymphocytic leukaemia, would interfere with a therapeutic strategy of T cell activation. Analysis of peripheral blood samples from patients prior and during fludarabine/cyclophosphamide therapy revealed rapid and sustained reduction of tumour cells but also of CD4+ and CD8+ T cells. This correlated with a significant cytotoxic activity of fludarabine/cyclophosphamide on T cells in vitro. Unexpectedly, T cells surviving fludarabine/cyclophosphamide treatment in vitro had a more mature phenotype, while fludarabine-treated T cells were significantly more responsive to mitogenic stimulation than their untreated counterparts and showed a shift towards TH1 cytokine secretion. In conclusion, fludarabine/cyclophosphamide therapy though inducing significant and relevant T cell depletion seems to generate a micromilieu suitable for subsequent T cell activation.

Phage Display Based Cloning of Proteins Interacting with the Cytoplasmic Tail of Membrane Immunoglobulins

The reduced quantity and quality of serum immunoglobulins (sIgs) in mutant mice expressing truncated cytoplasmic tails of IgE and IgG1 indicate an active role for the cytoplasmic domains of mIgG1 and mIgE. We used phage display technology to identify candidate proteins able to interact with the cytoplasmic tail of mIgE. Using a murine cDNA B cell library displayed on the surface of phage as prey and the 28 amino acid long cytoplasmic tail of IgE as bait, we isolated phage encoding the murine hematopoietic progenitor kinase 1 (HPK1). Surface plasmon resonance analysis measurements confirmed affinity of HPK1 to the mIgE cytoplasmic tail and revealed association to other immunoglobulin isotypes as well. Immunoprecipitation experiments, using lysates from two B cell lines expressing nitrophenyl (NP) specific mIgE molecules showed co-precipitation of IgE and HPK1. The interaction of HPK1 with the cytoplasmic domains of membrane immunoglobulins indicate an active role of the tails as part of an isotype specific signal transduction, independent from the Igα/Igβ heterodimers, and may represent a missing link to upstream regulatory elements of HPK1 activation.

Chronic lymphocytic leukaemia induces an exhausted T cell phenotype in the TCL1 transgenic mouse model

Although chronic lymphocytic leukaemia (CLL) is a B cell malignancy, earlier studies have indicated a role of T cells in tumour growth and disease progression. In particular, the functional silencing of antigen‐experienced T cells, called T cell exhaustion, has become implicated in immune evasion in CLL. In this study, we tested whether T cell exhaustion is recapitulated in the TCL1tg mouse model for CLL. We show that T cells express high levels of the inhibitory exhaustion markers programmed cell death 1 (PDCD1, also termed PD‐1) and lymphocyte‐activation gene 3 (LAG3), whereas CLL cells express high levels of CD274 (also termed PD‐ligand 1). In addition, the fraction of exhausted T cells increases with CLL progression. Finally, we demonstrate that exhausted T cells are reinvigorated towards CLL cytotoxicity by inhibition of PDCD1/CD274 interaction in vivo.

The IgE Antigen Receptor: A Key Regulator for the Production of IgE Antibodies

Immunoglobulins in general form a substantial component of serum proteins, and play a role in homeostatic mechanisms, a first line of defense against pathogenic organisms and in immunological memory. In the secreted form, immunoglobulins represent the effector arm of the humoral immune system. However, immunoglobulins are not only secreted, but can also be expressed on the surface of a B lymphocyte (membrane immunoglobulin), and, in this physical state, most likely convey signals to steer the B cell along its differentiation pathway. A step forward in the understanding of the role of membrane immunoglobulins other than membrane IgM or IgD was achieved with two mouse lines with mutations in the epsilon heavy chain gene. In IgEΔM1M2 mice serum IgE is reduced to less than 10% of normal mice, while IgEKVKΔtail mice show a reduction of 50%, reflecting a serious impairment of the IgE-mediated immune response. We think that the cytoplasmic tail of IgE is involved in a signal transduction which leads to the expression of high quantities and qualities of secreted IgE immunoglobulins.