Inga-Lill Mårtensson

Mutations in the homeobox gene HESX1/Hesx1 associated with septo-optic dysplasia in human and mouse

During early mouse development the homeobox gene Hesx1 is expressed in prospective forebrain tissue, but later becomes restricted to Rathkes pouch, the primordium of the anterior pituitary gland. Mice lacking Hesx1 exhibit variable anterior CNS defects and pituitary dysplasia. Mutants have a reduced prosencephalon, anopthalmia or micropthalmia, defective olfactory development and bifurcations in Rathkes pouch. Neonates exhibit abnormalities in the corpus callosum, the anterior and hippocampal commissures, and the septum pellucidum. A comparable and equally variable phenotype in humans is septo-optic dysplasia (SOD). We have cloned human HESX1 and screened for mutations in affected individuals. Two siblings with SOD were homozygous for an Arg53Cys missense mutation within the HESX1 homeodomain which destroyed its ability to bind target DNA. These data suggest an important role for Hesx1/HESX1 in forebrain, midline and pituitary development in mouse and human.

Complementary signaling through flt3 and interleukin-7 receptor alpha is indispensable for fetal and adult B cell genesis

Extensive studies of mice deficient in one or several cytokine receptors have failed to support an indispensable role of cytokines in development of multiple blood cell lineages. Whereas B1 B cells and Igs are sustained at normal levels throughout life of mice deficient in IL-7, IL-7Rα, common cytokine receptor gamma chain, or flt3 ligand (FL), we report here that adult mice double deficient in IL-7Rα and FL completely lack visible LNs, conventional IgM+ B cells, IgA+ plasma cells, and B1 cells, and consequently produce no Igs. All stages of committed B cell progenitors are undetectable in FL−/− × IL-7Rα−/− BM that also lacks expression of the B cell commitment factor Pax5 and its direct target genes. Furthermore, in contrast to IL-7Rα−/− mice, FL−/− × IL-7Rα−/− mice also lack mature B cells and detectable committed B cell progenitors during fetal development. Thus, signaling through the cytokine tyrosine kinase receptor flt3 and IL-7Rα are indispensable for fetal and adult B cell development.

VpreB1/VpreB2/λ5 Triple-Deficient Mice Show Impaired B Cell Development but Functional Allelic Exclusion of the IgH Locus

At the precursor B cell stage during bone marrow B cell development, Ig μH chain associates with surrogate L (SL) chain, which is encoded by the three genes VpreB1, VpreB2, and λ5, to form the pre-B cell receptor (pre-BCR). Surface expression of the pre-BCR is believed to signal both proliferation and allelic exclusion of the IgH locus. Mice which lack either VpreB1/VpreB2 or λ5 show a lack of precursor B cell expansion but normal IgH allelic exclusion. This would suggest that one of either λ5 or VpreB can make a pre-BCR-like complex which is still able to signal allelic exclusion but not proliferation. To investigate this, we established mice lacking all components of the SL chain. These mice showed severely impaired B cell development which was similar to that previously found in mice lacking either λ5 or VpreB1/VpreB2. Surprisingly, the IgH locus was still allelically excluded and thus the SL chain appears not to be involved in allelic exclusion.

OX40 Ligand and CD30 Ligand Are Expressed on Adult but Not Neonatal CD4+CD3− Inducer Cells: Evidence That IL-7 Signals Regulate CD30 Ligand but Not OX40 Ligand Expression

In this report, we have examined the expression of the T cell survival signals, OX40 ligand (OX40L) and CD30 ligand (CD30L) on CD4+CD3−CD11c−B220−IL-7Rα+ inducer cells from birth to adulthood in mice. We found that adult but not neonatal inducer cells expressed high levels of OX40L and CD30L, whereas their expression of TNF-related activation-induced cytokine (TRANCE) and receptor activator of NF-κB (RANK) was comparable. The failure of neonatal inducer cells to express the ligands that rescue T cells helps to explain why exposure to Ag in neonatal life induces tolerance rather than immunity. The expression of OX40L and CD30L on inducer cells increased gradually in the first few weeks of life achieving essentially normal levels around the time mice were weaned. We found that IL-7 signaling through the common cytokine receptor γ-chain was critical for the optimal expression of both TNF-related activation-induced cytokine and CD30L but not OX40L. Furthermore, glucocorticoids, which potently suppress T effector function, did not influence the expression of OX40L and CD30L in the presence of IL-7.

B-lineage commitment prior to surface expression of B220 and CD19 on hematopoietic progenitor cells

Commitment of hematopoietic progenitor cells to B-lymphoid cell fate has been suggested to coincide with the development of PAX5-expressing B220(+)CD19(+) pro-B cells. We have used a transgenic reporter mouse, expressing human CD25 under the control of the B-lineage-restricted Igll1 (lambda5) promoter to investigate the lineage potential of early progenitor cells in the bone marrow. This strategy allowed us to identify a reporter expressing LIN(-)B220(-)CD19(-)CD127(+)FLT3(+)SCA1(low)KIT(low) population that displays a lack of myeloid and a 90% reduction in in vitro T-cell potential compared with its reporter-negative counterpart. Gene expression analysis demonstrated that these lineage-restricted cells express B-lineage-associated genes to levels comparable with that observed in pro-B cells. These data suggest that B-lineage commitment can occur before the expression of B220 and CD19.

The pre-B cell receptor and its role in proliferation and Ig heavy chain allelic exclusion

The pre-B cell receptor (pre-BCR) is composed of the immunoglobulin (Ig) heavy (microH) chain and the surrogate light chain encoded by VpreB and lambda5. The pre-BCR has been implicated in precursor B cell proliferation, differentiation and IgH chain allelic exclusion. B cell development in mice lacking the transmembrane form of microH chain is blocked at the precursor B cell stage: the cells cannot proliferate or differentiate further and the IgH locus is allelically included. In mice lacking lambda5, the precursor B cells, although unable to proliferate, can nonetheless differentiate, whereas the IgH locus is allelically excluded. It was, therefore, postulated that microH chain together with VpreB could form a pre-BCR-like receptor that would allow IgH allelic exclusion but not proliferation. In mice lacking both VpreB genes, precursor B cells do not proliferate but are able to differentiate. Surprisingly, the IgH locus is allelically excluded. This suggests that microH chains find other partner proteins to signal allelic exclusion.

Surface μ Heavy Chain Signals Down-Regulation of the V(D)J-Recombinase Machinery in the Absence of Surrogate Light Chain Components

Early B cell development is characterized by stepwise, ordered rearrangement of the immunoglobulin (Ig) heavy (HC) and light (LC) chain genes. Only one of the two alleles of these genes is used to produce a receptor, a phenomenon referred to as allelic exclusion. It has been suggested that pre–B cell receptor (pre-BCR) signals are responsible for down-regulation of the VDJH-recombinase machinery (Rag1, Rag2, and terminal deoxynucleotidyl transferase [TdT]), thereby preventing further rearrangement on the second HC allele. Using a mouse model, we show that expression of an inducible μHC transgene in Rag2−/− pro–B cells induces down-regulation of the following: (a) TdT protein, (b) a transgenic green fluorescent protein reporter reflecting endogenous Rag2 expression, and (c) Rag1 primary transcripts. Similar effects were also observed in the absence of surrogate LC (SLC) components, but not in the absence of the signaling subunit Ig-α. Furthermore, in wild-type mice and in mice lacking either λ5, VpreB1/2, or the entire SLC, the TdT protein is down-regulated in μHC+LC− pre–B cells. Surprisingly, μHC without LC is expressed on the surface of pro–/pre–B cells from λ5−/−, VpreB1−/−VpreB2−/−, and SLC−/− mice. Thus, SLC or LC is not required for μHC cell surface expression and signaling in these cells. Therefore, these findings offer an explanation for the occurrence of HC allelic exclusion in mice lacking SLC components.

The pre‐B‐cell receptor induces silencing of VpreB and λ5 transcription

The pre‐B‐cell receptor (pre‐BCR), composed of Ig heavy and surrogate light chain (SLC), signals pre‐BII‐cell proliferative expansion. We have investigated whether the pre‐BCR also signals downregulation of the SLC genes (VpreB and λ5), thereby limiting this expansion. We demonstrate that, as BM cells progress from the pre‐BI to large pre‐BII‐cell stage, there is a shift from bi‐ to mono‐allelic λ5 transcription, while the second allele is silenced in small pre‐BII cells. A VpreB1‐promoter‐driven transgene shows the same pattern, therefore suggesting that VpreB1 is similarly regulated and thereby defines the promoter as a target for transcriptional silencing. Analyses of pre‐BCR‐deficient mice show a temporal delay in λ5 downregulation, thereby demonstrating that the pre‐BCR is essential for monoallelic silencing at the large pre‐BII‐cell stage. Our data also suggest that SLP‐65 is one of the signaling components important for this process. Furthermore, the VpreB1/λ5 alleles undergo dynamic changes with respect to nuclear positioning and heterochromatin association, thereby providing a possible mechanism for their transcriptional silencing.

Two Pathways of B‐Lymphocyte Development in Mouse Bone Marrow and the Roles of Surrogate L Chain in this Development

The commitment of pluripotent stem cells and lymphoid progenitors to the Blineage pathway of differentiation in bone marrow of mice has been investigated in many laboratories. One aim of such studies is to analyze the molecular steps and cellular stages in which B-lineage progenitors and precursors rearrange immunoglobulin (Ig) heavy (H) and light (L) chain gene segments (V, D, J) to express first H, then L chains, and thereby generate a potential repertoire of B lymphocytes. The selection of these cells into a repertoire available in the peripheral pool of mature B cells has also been extensively examined (for reviews see Osmond 1991, Tsubata & Nishikawa 1991, Hardy 1992, Rajewsky 1992, Chen & Alt 1993, Rolink & Melchers 1993a, b). Fig. 1 summarizes some of our current knowledge of the size and cell cycle status of the major B-lineage precursor subpopulations in bone marrow, their expression of markers, and their potential for in vitro proliferation and for in vivo transplantability. It describes two pathways of differentiation, one of which (pathway 1) selects for productively H chain-gene-rearranged, fiH chain-expressing precursors, while the other pathway (2) appears to occur without selection for products of Ig gene rearrangements, and will proceed via a cellular program of surface marker and functional changes even without Ig-gene rearrangements. These two pathways will be explained in more detail. In this review article we summarize our recent work which is concerned with

The pre-B cell receptor checkpoint

B lymphocytes are essential antibody‐producing cells of the immune system. During the development of progenitor B cells to mature B cells that express a membrane‐bound antibody, the B cell receptor (BCR), the cells undergo selection at several checkpoints, which ensures that a diverse antibody repertoire is generated and that the BCRs recognise foreign‐, but not self‐, antigens. In this review, we consider the pre‐BCR checkpoint. Mutations or alterations that affect this checkpoint underpin the development of pre‐B cell leukemias, primary immunodeficiency, and possibly, systemic autoimmunity.