The murine homologue of the previously identified human "pre-B-cell colony-enhancing factor" (PBEF) gene coding for a putative cytokine has been identified by screening a subtractive library enriched in genes expressed in activated T lymphocytes. Unlike most cytokine genes known to date, the PBEF gene is ubiquitously expressed in lymphoid and non-lymphoid tissues and displays significanthomology with genes from primitive metazoans (marine sponges) and prokaryotic organisms. Recently, a bacterial protein encoded by nadV, a gene from the prokaryote Haemophilus ducreyi displaying significant homology with PBEF, has been identified as a nicotinamide phosphoribosyltranferase (NAmPRTase), an enzyme involved in nicotinamide adenine dinucleotide (NAD) biosynthesis.Using a panel of antibodies to murine PBEF, we demonstrate in this work that, similarly to its microbial counterpart, the murine protein is a NAmPRTase, catalyzing the condensation of nicotinamide with 5-phosphoribosyl-1-pyrophosphate to yield nicotinamide mononucleotide, an intermediate in the biosynthesis of NAD. The role of PBEF as a NAmPRTase was further confirmed by showing that the mouse gene was able to confer the ability to grow in the absence of NAD to a NAmPRTase-defective bacterial strain. The present findings are in keeping with the ubiquitous nature of this protein, and indicatethat NAD biosynthesis may play an important role in lymphocyte activation.
The aim of this study was to test whether the nature of the antigen-presenting cell (APC) can influence the Th1 / Th2 balance in vivo. Our data show that dendritic cells (DC), pulsed extra corporeally with antigen, induced the development of cells secreting IL-2, IFN-γ and IL-4 upon antigen rechallenge in vitro. Priming with peritoneal macrophages sensitized cells that produced IL-4 but not IFN-γ. To identify the factors involved in T helper development, mice were primed with APC with or without treatment with neutralizing antibodies to co-stimulatory molecules or cytokines. Our results indicate that priming with DC or macrophages is strictly dependent on the CD28-CTLA4/ B7 interaction. Of note, CD86 provides the initial signal to induce naive T cells to become IL-4 producers, whereas CD80 is a more neutral differentiation signal. IL-12, released by the DC, appears as a potent and obligatory inducer of differentiation for IFN-γ-producing cells. IL-6, although produced by both APC populations, is necessary to direct activation of the Th2-type response by macrophages but not by DC.
The potent accessory properties of dendritic cells (DC) develop sequentially during a process termed "maturation." Splenic DC undergo functional maturation in vivo in response to the bacterial product LPS and migrate from the marginal zone to the T cell areas. The redistribution of fully mature DC, which present Ags encountered in the periphery, in the T cell area is likely to result in T cell priming. Unexpectedly, we found that DC rapidly die by apoptosis once they have entered the T cell zone. Injection of OVA peptide in OVA-specific, TCR-transgenic mice strongly delays the LPS-induced apoptosis of DC in situ. We conclude that mature DC are programmed to die unless they receive a survival signal from T cells and that the regulation of DC survival may be a mechanism aimed at controlling the initiation and the termination of the immune response.
We have found that syngeneic Ab2s in the antiarsonate system are serologically and structurally similar to one another. In contrast, the allogeneic Ab2 response is heterogeneous and derives from a large number of unrelated germline gene segments. The Ab2 response of the BALB/c strain to polyclonal A/J Ars A molecules can probably best be compared with a response to a foreign protein and might have been predicted in a strain that completely lacks the H chain V region gene from which the Ab1 derives. Partial variable region sequences of Ab2s from three other systems in addition to previously reported Ab2 structures indicates that this difference in allogeneic vs. syngeneic Ab2s may be a general phenomena. These data support Jerne's hypothesis of complementary V region genes existing in the germline. However, there is good evidence that these antiidiotypic antibodies are not derived directly from the germline, as somatic processes most likely play an important role in their generation. The D segments of Ab2s in the arsonate system as well as in other systems, are novel in structure and cannot easily be explained by previously described germline D segments. D-D fusion may play a role in the generation of the third hypervariable region in these antibodies.
Presentation of antigen to T lymphocytes without the appropriate co-stlmulatory signals results in a state of antigen-specific unresponsiveness. Despite the presumed importance of the B7-CD28 interaction for the initiation and maintenance of T cell-mediated immune responses, relatively few studies have addressed the regulation of B7 expression. We have studied the expression of the CD80 (B7-1) and B7-2 molecules on peripheral blood monocytes following different activation signals, and it was demonstrated that not only IFN-γ, but also granulocyte macrophage colony stimulating factor can induce CD80 expression on monocytes. In addition, we found that cross linking of FcR on monocytes strongly inhibits the up-regulation of CD80 and B7-2, with as a functional consequence that the capacity to function as antigen presenting cells (APC) and to.stimulate T cell activation is severely impaired. When cultures were prepared In 96-well plates coated with human IgG, stimulation of T cells with allogenelc monocytes resulted only in modest T cell proliferation and no detectable IL-2 secretion as compared with untreated culture plates or plates coated with Fab fragments of human IgG. Under these conditions cross-linking of CD28 on the T cells with specific mAb completely reverted the inhibitory effect observed after culture on IgG-coated plates. Furthermore, FcR cross-linking on monocytes strongly Inhibited the capacity of monocytes to Induce a specific memory T cell response to viral, bacterial and fungal antigens, whereas the treatment did not impair the capacity of the T cells to respond to pokeweed mitogen, phytohemagglutinin and concanavalin A. We conclude that after FcR cross-linking, the impaired APC function is most likely due to the inability of monocytes to provide the essential costimulatory signals to the T cells via the B7–CD28/CTLA-4 interaction.
Glucocorticoids (GC) are widely used anti-inflammatory agents known to suppress T cell activation by interfering with the TCR activation cascade. The attenuation of early TCR signaling events by these compounds has been recently attributed to a selective displacement of key signaling proteins from membrane lipid rafts. In this study, we demonstrate that GC displace the acyl-bound adaptor proteins linker for activation of T cells and phosphoprotein associated with glycosphingolipid-enriched microdomains from lipid rafts of murine T cell hybridomas, possibly by inhibiting their palmitoylation status. Analysis of the lipid content of the membrane rafts revealed that GC treatment led to a significant decrease in palmitic acid content. Moreover, we found an overall decrease in the proportion of raft-associated saturated fatty acids. These changes were consistent with a decrease in fluorescence anisotropy of isolated lipid rafts, indicating an increase in their fluidity. These findings identify the mechanisms underlying the complex inhibitory effects of glucocorticoids on early TCR signaling and suggest that some of the inhibitory properties of GC on T cell responses may be related to their ability to affect the membrane lipid composition and the palmitoylation status of important signaling molecules.
We present a statistical method for detection of palindromes in mRNA or DNA, starting from the protein sequence. Analysis of immunoglobulin genes by this method demonstrates that palindromic sequences are not randomly distributed. They are located at each side of the hypervariable regions in the variable (V) genes, whereas no such regular design is observed in the constant (C) genes. In addition, palindromic sequences overlap the V-C junction in all immunoglobulin classes and significant palindromes are present near residue 216 of the heavy chain, which is the end of deletions in many heavy chain diseases. The relevance of these palindromes to gene translocation and generation of diversity in antibodies is discussed.
