Human T cell differentiation : basic aspects and their clinical applications

Immune recognition plays a central role in our understanding of the function of the immune system. The ability to specifically recognize foreign antigens allows selective but efficient actions of the immune system against all kinds of pathogens. This is mediated by antigen-specific receptors on B and T lymphocytes. Immunoglobulin (lg) molecules represent the antigenspecific receptors of B lymphocytes, while the T cell receptor (TeA) has this function in T lymphocytes (1). Although these two types of antigen receptors have remarkable similarities in protein structure and their encoding genes, they differ significantly in their ability to interact with antigens (1). Via their surface membrane lg (Smlg) molecules, B lymphocytes are able to recognize antigens in their native configuration either free in solution, on surfaces or on cell membranes (1). TeA molecules ofT lymphocytes can only recognize processed or degraded antigens which are physically associated with major histocompatibility complex (MHC) molecules (2,3). This TeA-mediated recognition is therefore called MHC-restricted antigen recognition (2,3). Expression of Smlg or TcA molecules by lymphocytes is acquired during lymphoid differentiation via several rearrangement processes in the lg or TcR genes (3-7). B lymphopoiesis mainly occurs in the bone marrow (8), while the thymus is thought to represent the main tissue compartment for T lymphopoiesis (9-11 ). During T cell differentiation in the thymus the T lymphocytes are "educated" for their future functions, i.e. T cells which recognize self antigens are eliminated (negative selection), while positive selection occurs for T cells which recognize foreign (non-self) antigens in association with self-MHC molecules (12-14). Upon recognition of a TeA-compatible antigen, T lymphocytes are activated, start to proliferate and exhibit their regulatory or cytotoxic functions (2). These T cell functions play a central role in the regulation of the immune system. The T lymphocytes probably coordinate immune processes via cellular interactions and lymphokines and in this way adjust and harmonize the actions of the immune system. The TeA consists of two chains, which are closely associated with the CD3 protein complex (TcA-CD3). Th

Replication history of B lymphocytes reveals homeostatic proliferation and extensive antigen-induced B cell expansion

The contribution of proliferation to B lymphocyte homeostasis and antigen responses is largely unknown. We quantified the replication history of mouse and human B lymphocyte subsets by calculating the ratio between genomic coding joints and signal joints on kappa-deleting recombination excision circles (KREC) of the IGK-deleting rearrangement. This approach was validated with in vitro proliferation studies. We demonstrate that naive mature B lymphocytes, but not transitional B lymphocytes, undergo in vivo homeostatic proliferation in the absence of somatic mutations in the periphery. T cell–dependent B cell proliferation was substantially higher and showed higher frequencies of somatic hypermutation than T cell–independent responses, fitting with the robustness and high affinity of T cell–dependent antibody responses. More extensive proliferation and somatic hypermutation in antigen-experienced B lymphocytes from human adults compared to children indicated consecutive responses upon additional antigen exposures. Our combined observations unravel the contribution of proliferation to both B lymphocyte homeostasis and antigen-induced B cell expansion. We propose an important role for both processes in humoral immunity. These new insights will support the understanding of peripheral B cell regeneration after hematopoietic stem cell transplantation or B cell–directed antibody therapy, and the identification of defects in homeostatic or antigen-induced B cell proliferation in patients with common variable immunodeficiency or another antibody deficiency

Molecular Monitoring of Lymphoma

This chapter provides the background information of the PCR targets for molecular MRD monitoring (i.e., Ig/TCR gene rearrangements and chromosome aberrations), explains how these targets can be identified. [...

Molecular Monitoring of Lymphoma

This chapter provides the background information of the PCR targets for molecular MRD monitoring (i.e., Ig/TCR gene rearrangements and chromosome aberrations), explains how these targets can be identified. [...

PID Comes Full Circle: Applications of V(D)J Recombination Excision Circles in Research, Diagnostics and Newborn Screening of Primary Immunodeficiency Disorders

The vast majority of patients suffering from a primary immunodeficiency (PID) have defects in their T- and/or B-cell compartments. Despite advances in molecular diagnostics, in many patients no underlying genetic defect has been identified. B- and T-lymphocytes are unique in their ability to create a receptor by genomic rearrangement of their antigen receptor genes via V(D)J recombination. During this process, stable circular excision products are formed that do not replicate when the cell proliferates. Excision circles can be reliably quantified using real-time quantitative (RQ-)PCR techniques. Frequently occurring δREC–ψJα T-cell receptor excision circles (TRECs) have been used to assess thymic output and intronRSS–Kde recombination excision circles (KREC) to quantify B-cell replication history. In this perspective, we describe how TRECs and KRECs are formed during precursor – T- and B-cell differentiation, respectively. Furthermore, we discuss new insights obtained with TRECs and KRECs and specifically how these excision circles can be applied to support therapy monitoring, patient classification and newborn screening of PID

Basic helix-loop-helix proteins E2A and HEB induce immature T-cell receptor rearrangements in nonlymphoid cells

T-cell receptor (TCR) gene rearrangements are mediated via V(D)J recombination, which is strictly regulated during lymphoid differentiation, most probably through the action of specific transcription factors. Investigated was whether cotransfection of RAG1 and RAG2 genes in combination with lymphoid transcription factors can induce TCR gene rearrangements in nonlymphoid human cells. Transfection experiments showed that basic helix-loop-helix transcription factors E2A and HEB induce rearrangements in the TCRD locus (Ddelta2-Ddelta3 and Vdelta2-Ddelta3) and TCRG locus (psi Vgamma7-Jgamma2.3 and Vgamma8-Jgamma2.3). Analysis of these rearrangements and their circular excision products revealed some peculiar characteristics. The Vdelta2-Ddelta3 rearrangements were formed by direct coupling without intermediate Ddelta2 gene segment usage, and most Ddelta2-Ddelta3 recombinations occurred via direct coupling of the respective upstream and downstream recombination signal sequences (RSSs) with deletion of the Ddelta2 and Ddelta3 coding sequences. Subsequently, the E2A/HEB-induced TCR gene recombination patterns were compared with those in early thymocytes and acute lymphoblastic leukemias of T- and B-lineage origin, and it was found that the TCR rearrangements in the transfectants were early (immature) and not necessarily T-lineage specific. Apparently, some parts

Comparative analysis of Ig and TCR gene rearrangements at diagnosis and at elapse of childhood precursor-B–ALL provides improved strategies for selection of stable PCR targets for monitoring of minimal residual disease

Immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements are excellent patient-specific polymerase chain reaction (PCR) targets for detection of minimal residual disease (MRD) in acute lymphoblastic leukemia (ALL), but they might be unstable during the disease course. Therefore, we performed detailed molecula

Involvement of Artemis in nonhomologous end-joining during immunoglobulin class switch recombination

DNA double-strand breaks (DSBs) introduced in the switch (S) regions are intermediates during immunoglobulin class switch recombination (CSR). These breaks are subsequently recognized, processed, and joined, leading to recombination of the two S regions. Nonhomologous end-joining (NHEJ) is believed to be the principle mechanism involved in DSB repair during CSR. One important component in NHEJ, Artemis, has however been considered to be dispensable for efficient CSR. In this study, we have characterized the S recombinational junctions from Artemis-deficient human B cells. Sμ–Sα junctions could be amplified from all patients tested and were characterized by a complete lack of “direct” end-joining and a remarkable shift in the use of an alternative, microhomology-based end-joining pathway. Sμ–Sγ junctions could only be amplified from one patient who carries “hypomorphic” mutations. Although these Sμ–Sγ junctions appear to be normal, a significant increase of an unusual type of sequential switching from immunoglobulin (Ig)M, through one IgG subclass, to a different IgG subclass was observed, and the Sγ–Sγ junctions showed long microhomologies. Thus, when the function of Artemis is impaired, varying modes of CSR junction resolution may be used for different S regions. Our findings strongly link Artemis to the predominant NHEJ pathway during CSR