B cell depletion in autoimmune disease

The CD20 cell marker appears early in the process of B cell development. In this review we focus on the results of attempts to utilize B cell depletion based on the use of a chimeric monoclonal antibody (MAb) specific for human CD20, rituximab, for the treatment of patients with autoimmune diseases. In 1997, rituximab was approved for the treatment of low-grade B cell non-Hodgkin's lymphoma. Following these encouraging results, rituximab started to be used experimentally in other diseases presumed to be due to B cell pathology. The first autoimmune disease to be treated effectively was chronic idiopathic thrombocytopaenia. More recent success has been demonstrated in patients with rheumatoid arthritis and systemic lupus erythematosus.     

Cell-autonomous and cell non-autonomous signaling through endothelin receptor B during melanocyte development

The endothelin receptor B gene (Ednrb) encodes a G-protein-coupled receptor that is expressed in a variety of cell types and is specifically required for the development of neural crest-derived melanocytes and enteric ganglia. In humans, mutations in this gene are associated with Waardenburg-Shah syndrome, a disorder characterized by pigmentation defects, deafness and megacolon. To address the question of whether melanocyte development depends entirely on a cell-autonomous action of Ednrb, we performed a series of tissue recombination experiments in vitro, using neural crest cell cultures from mouse embryos carrying a novel Ednrb-null allele characterized by the insertion of a lacZ marker gene. The results show that Ednrb is not required for the generation of early neural crest-derived melanoblasts but is required for the expression of the differentiation marker tyrosinase. Tyrosinase expression can be rescued, however, by the addition of Ednrb wild-type neural tubes. These Ednrb wild-type neural tubes need not be capable of generating melanocytes themselves, but must be capable of providing KIT ligand, the cognate ligand for the tyrosine kinase receptor KIT. In fact, soluble KIT ligand is sufficient to induce tyrosinase expression in Ednrb-deficient cultures. Nevertheless, these tyrosinase-expressing, Ednrb-deficient cells do not develop to terminally differentiated, pigmented melanocytes. Pigmentation can be induced, however, by treatment with tetradecanoyl phorbol acetate, which mimics EDNRB signaling, but not by treatment with endothelin 1, which stimulates the paralogous receptor EDNRA. The results suggest that Ednrb plays a significant role during melanocyte differentiation and effects melanocyte development by both cell non-autonomous and cell-autonomous signaling mechanisms.     

Ski and SnoN: negative regulators of TGF-beta signaling

Ski and SnoN are unique proto-oncoproteins in that they can induce both oncogenic transformation and terminal muscle differentiation when expressed at high levels. Recent studies using in vitro and in vivo approaches have begun to unravel the complex roles of Ski and SnoN in tumorigenesis and embryonic development. The identification of Ski and SnoN as important negative regulators of signal transduction by the transforming growth factor-beta superfamily of cytokines provides a valuable molecular basis for the complex functions of Ski and SnoN.     

Type-A Arabidopsis response regulators are partially redundant negative regulators of cytokinin signaling

Type-A Arabidopsis (Arabidopsis thaliana) response regulators (ARRs) are a family of 10 genes that are rapidly induced by cytokinin and are highly similar to bacterial two-component response regulators. We have isolated T-DNA insertions in six of the type-A ARRs and constructed multiple insertional mutants, including the arr3,4,5,6,8,9 hextuple mutant. Single arr mutants were indistinguishable from the wild type in various cytokinin assays; double and higher order arr mutants showed progressively increasing sensitivity to cytokinin, indicating functional overlap among type-A ARRs and that these genes act as negative regulators of cytokinin responses. The induction of cytokinin primary response genes was amplified in arr mutants, indicating that the primary response to cytokinin is affected. Spatial patterns of ARR gene expression were consistent with partially redundant function of these genes in cytokinin signaling. The arr mutants show altered red light sensitivity, suggesting a general involvement of type-A ARRs in light signal transduction. Further, morphological phenotypes of some arr mutants suggest complex regulatory interactions and gene-specific functions among family members.     

Multiple positive and negative regulators of signaling by the EGF-receptor

Signaling via the epidermal growth factor (EGF)-receptor family is subject to regulation and modulation by multiple ligands, effectors and negative regulators, as well as regulation by heterodimerization between family members and crosstalk between heterologous signaling pathways. Besides serving as a paradigm for receptor tyrosine kinases in general, this family is crucial for development and is often mutated or amplified in human tumors.     

Leupaxin negatively regulates B cell receptor signaling

The role of the paxillin superfamily of adaptor proteins in B cell antigen receptor (BCR) signaling has not been studied previously. We show here that leupaxin (LPXN), a member of this family, was tyrosine-phosphorylated and recruited to the plasma membrane of human BJAB lymphoma cells upon BCR stimulation and that it interacted with Lyn (a critical Src family tyrosine kinase in BCR signaling) in a BCR-induced manner. LPXN contains four leucine-rich sequences termed LD motifs, and serial truncation and specific domain deletion of LPXN indicated that its LD3 domain is involved in the binding of Lyn. Of a total of 11 tyrosine sites in LPXN, we mutated Tyr(22), Tyr(72), Tyr(198), and Tyr(257) to phenylalanine and demonstrated that LPXN was phosphorylated by Lyn only at Tyr(72) and that this tyrosine site is proximal to the LD3 domain. The overexpression of LPXN in mouse A20 B lymphoma cells led to the suppression of BCR-induced activation of JNK, p38 MAPK, and, to a lesser extent, Akt, but not ERK and NFkappaB, suggesting that LPXN can selectively repress BCR signaling. We further show that LPXN suppressed the secretion of interleukin-2 by BCR-activated A20 B cells and that this inhibition was abrogated in the Y72F LPXN mutant, indicating that the phosphorylation of Tyr(72) is critical for the biological function of LPXN. Thus, LPXN plays an inhibitory role in BCR signaling and B cell function.     

Planar cell polarity regulators in asymmetric organogenesis during development and disease

The phenomenon of planar cell polarity is critically required for a myriad of morphogenetic processes in metazoan and is accurately controlled by several conserved modules. Six “core” proteins, including Frizzled, Flamingo (Celsr), Van Gogh (Vangl), Dishevelled, Prickle, and Diego (Ankrd6), are major components of the Wnt/planar cell polarity pathway. The Fat/Dchs protocadherins and the Scrib polarity complex also function to instruct cellular polarization. In vertebrates, all these pathways are essential for tissue and organ morphogenesis, such as neural tube closure, left–right symmetry breaking, heart and gut morphogenesis, lung and kidney branching, stereociliary bundle orientation, and proximal–distal limb elongation. Mutations in planar polarity genes are closely linked to various congenital diseases. Striking advances have been made in deciphering their contribution to the establishment of spatially oriented pattern in developing organs and the maintenance of tissue homeostasis. The challenge remains to clarify the complex interplay of different polarity pathways in organogenesis and the link of cell polarity to cell fate specification. Interdisciplinary approaches are also important to understand the roles of mechanical forces in coupling cellular polarization and differentiation. This review outlines current advances on planar polarity regulators in asymmetric organ formation, with the aim to identify questions that deserve further investigation.     

Protein tyrosine phosphatases as negative regulators of mitogenic signaling

The regulation of tyrosine phosphorylation represents a key mechanism governing cell proliferation. In fibroblasts, inputs from both growth factor and extracellular matrix receptors are required for cell division. Triggering such receptors induces a wave of tyrosine phosphorylation on key signaling molecules, culminating in the activation of cyclin-dependent kinases and cell cycle progression. In general, protein tyrosine kinases stimulate, while protein tyrosine phosphatases inhibit, such cell proliferation pathways. The role of protein tyrosine kinases in mitogenesis has been extensively studied, but the identity and targets of the protein tyrosine phosphatases that regulate cell growth are not well described. In this review, I will survey recent advances in the identification and regulation of protein tyrosine phosphatases that downregulate cell proliferation. J. Cell. Physiol. 180:173–181, 1999. © 1999 Wiley-Liss, Inc.     

Type IV collagens and Dpp positive and negative regulators of signaling

Decapentaplegic (Dpp) is an essential morphogen in the TGF-β/BMP superfamily which patterns fields of cells during multiple stages of Drosophila development, including the ovary and embryo. We have found that type IV collagens bind to Dpp and play essential roles in the regulation of its signaling during these two developmental stages. This article primarily focuses on type IV collagens and embryonic Dpp signaling to discuss aspects of the type IV collagen mutant phenotype in the context of additional data from the field. In addition, the restriction of Dpp signaling in the the ovary by type IV collagens is described, as the differences between the embryonic and ovarian Dpp sources result in distinct effects of collagen IV proteins in the two systems.     

Genetic and transformation studies reveal negative regulation of ERS1 ethylene receptor signaling in Arabidopsis

Background  

Ethylene receptor single mutants of Arabidopsis do not display a visibly prominent phenotype, but mutants defective in multiple ethylene receptors exhibit a constitutive ethylene response phenotype. It is inferred that ethylene responses in Arabidopsis are negatively regulated by five functionally redundant ethylene receptors. However, genetic redundancy limits further study of individual receptors and possible receptor interactions. Here, we examined the ethylene response phenotype in two quadruple receptor knockout mutants, (ETR1) ers1 etr2 ein4 ers2 and (ERS1) etr1 etr2 ein4 ers2, to unravel the functions of ETR1 and ERS1. Their functions were also reciprocally inferred from phenotypes of mutants lacking ETR1 or ERS1. Receptor protein levels are correlated with receptor gene expression. Expression levels of the remaining wild-type receptor genes were examined to estimate the receptor amount in each receptor mutant, and to evaluate if effects of ers1 mutations on the ethylene response phenotype were due to receptor functional compensation. As ers1 and ers2 are in the Wassilewskija (Ws) ecotype and etr1, etr2, and ein4 are in the Columbia (Col-0) ecotype, possible effects of ecotype mixture on ethylene responses were also investigated.     

Polyclonal B cell activation by the Eta-1 cytokine and the development of systemic autoimmune disease

Studies of systemic autoimmune disease have led to the view that initiation and progression of the disease process reflects chronic and sustained B cell activation by unidentified polyclonal activating agents. In earlier studies, we found that T cells from MRL/1 mice, which develop murine lupus, express very high levels of a newly defined T cell cytokine, Eta-1. Inasmuch as chronic and sustained B cell stimulation by T cells is a cardinal feature of MRL/1 disease, we determined the effects of this cytokine on Ig production by B cells. We show that both recombinant and biochemically purified natural Eta-1 stimulate IgM and IgG production by mixtures of B cells and macrophages from the autoimmune MRL/l strain. Additional studies suggest that optimal Ig production by Eta-1 may require macrophages and reflect enhanced Ig production by large B cells. These findings support the view that elevated levels of endogenous Eta-1 may cause chronic and sustained polyclonal B cell activation that leads to autoimmune disease in this murine model.     

Level of B cell antigen receptor surface expression influences both positive and negative selection of B cells during primary development

To examine the effect of B cell Ag receptor (BCR) surface density on B cell development, we studied multiple lines of mice containing various copy numbers of an IgH micro delta transgene. The V(H) gene in this transgene encodes multireactive BCRs with low affinity for self Ags. These BCRs promote differentiation to a B cell subpopulation that shares some, but not all of the properties of marginal zone (MZ) B cells. Surface BCR level was found to be related to transgene gene copy number in these mice. In mice containing 1-15 copies of the transgene, elevated surface BCR levels were correlated with increased numbers of B cells in the MZ-like subset. However, in mice containing 20-30 copies of the transgene, massive clonal deletion of B cells was observed in the bone marrow, few B cells populated the spleen, and B cells were essentially absent from the lymph nodes. These data support the idea that autoantigens mediate not only negative, but positive selection of developing B cells as well. More importantly, they illustrate the profound influence of BCR surface density on the extent to which either of these selective processes take place.