Idiotypes of anti-Ia antibodies. I. Expression of the 14-4-4S idiotype in humoral immune responses.

The idiotype of a mouse monoclonal anti-I-E antibody, 14-4-4S, has been studied using a heterologous anti-idiotypic reagent. This antibody recognizes Ia. 7, an antigenic specificity present in all strains expressing a product of the I-E subregion. Expression of the 14-4-4S idiotype in humoral immune responses was analyzed by an idiotype-specific enzyme-linked immunosorbent assay system. The idiotype was readily detectable in C3H.SW anti-C3H alloantisera, the same immunization combination from which the hybridoma was derived. Absorption analysis demonstrated the anti-I-E specificity of the idiotype-positive molecules in these alloantisera. Penetrance of idiotype expression was high among individual C3H.SW immune mice (9 of 10 tested). To examine genetic requirements for idiotype expression, an immunization was performed using as responders CWB mice, congenic with C3H.SW but differing at the heavy chain allotype loci. Immune sera of individual CWB mice contained very little or no idiotype, demonstrating that levels of idiotype expression are influenced by allotype-linked genes, although the influence of other genes has not been ruled. The 14-4-4S idiotype therefore represents a shared idiotype of anti-Ia antibodies and provides opportunities for analysis of the idiotypes of cellular receptors for the corresponding Ia antigen

Involvement of Mhc Loci in immune responses that are not Ir-gene-controlled

Twenty-nine randomly chosen, soluble antigens, many of them highly complex, were used to immunize mice of two strains, C3H and B10.RIII. Lymphnode cells from the immunized mice were restimulated in vitro with the priming antigens and the proliferative response of the cells was determined. Both strains were responders to 28 of 29 antigens. Eight antigens were then used to immunize 11 congenic strains carrying different H-2 haplotypes, and the T-cell proliferative responses of these strains were determined. Again, all the strains responded to seven of the eight antigens. These experiments were then repeated, but this time -antibodies specific for the A (AA) or E (EE) molecules were added to the culture to block the in vitro responsiveness. In all but one of the responses, inhibition with both A-specific and E-specific antibodies was observed. The response to one antigen (Blastoinyces) was exceptional in that some strains were nonresponders to this antigen. Furthermore, the response in the responder strains was blocked with A-specific, but not with E-specific, antibodies. The study demonstrates that responses to antigens not controlled by Irr genes nevertheless require participation of class II Mhc molecules. In contrast to Ir gene-controlled responses involving either the A- or the E-molecule controlling loci (but never both), the responses not Ir-controlled involve participation of both A- and E-controlling loci. The lack of Ir-gene control is probably the result of complexity of the responses to multiple determinants. There is thus no principal difference between responses controlled and those not controlled by Ir genes: both types involve the recognition of the antigen, in the context of Mhc molecules

Propanil Exposure Induces Delayed but Sustained Abrogation of Cell-Mediated Immunity through Direct Interference with Cytotoxic T-Lymphocyte Effectors

The postemergent herbicide propanil (PRN; also known as 3,4-dichloropropionanilide) is used on rice and wheat crops and has well-known immunotoxic effects on various compartments of the immune system, including T-helper lymphocytes, B lymphocytes, and macrophages. It is unclear, however, whether PRN also adversely affects cytotoxic T lymphocytes (CTLs), the primary (1°) effectors of cell-mediated immunity. In this study we examined both the direct and indirect effects of PRN exposure on CTL activation and effector cell function to gauge its likely impact on cell-mediated immunity. Initial experiments addressed whether PRN alters the class I major histocompatibility complex (MHC) pathway for antigen processing and presentation by antigen-presenting cells (APCs), thereby indirectly affecting effector function. These experiments demonstrated that PRN does not impair the activation of CTLs by PRN-treated APCs. Subsequent experiments addressed whether PRN treatment of CTLs directly inhibits their activation and revealed that 1° alloreactive CTLs exposed to PRN are unimpaired in their proliferative response and only marginally inhibited in their lytic activity. Surprisingly, secondary stimulation of these alloreactive CTL effectors, however, even in the absence of further PRN exposure, resulted in complete abrogation of CTL lytic function and a delayed but significant long-term effect on CTL responsiveness. These findings may have important implications for the diagnosis and clinical management of anomalies of cell-mediated immunity resulting from environmental exposure to various herbicides and other pesticides

STRAIN DIFFERENCES IN THE EXPRESSION OF THE EPA-1-RESTRICTING ELEMENT

Epa-1-specific cytotoxic T lymphocytes (CTL) lyse epidermal cells (EC) of different Epa-1 + H-2 k strains, such as AKR, CBA, C58, and RF, at different levels. We used an H-2K k -specific monoclonal antibody (mAb) to test the hypothesis that this phenomenon is due to differences in the H-2-restricting element. Initially, we established the specificity of this mAb for the Epa-1-restricting element by demonstrating its capacity to inhibit the lysis of CBA EC by Epa-1-specific CTL. We then used it as the probe in a cellular radioimmunoassay to quantify the expression of the restricting element by EC of different H-2 k strains. We found that C58 and RF EC bound significantly less of the mAb than did CBA EC. Although AKR also bound less of the mAb than did CBA EC, the difference was not statistically significant. To examine the generality of this phenomenon, we quantified the expression of K k antigens on spleen cells (SC) of the same four strains. We found that RF SC, but not AKR or C58 SC, bound significantly less of the K k mAb than did CBA SC. Thus, the differential CTL lysis of Epa-1 + EC of different strains probably reflects differences in expression of the H-2-restricting element rather than of the nominal antigen.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/75485/1/j.1744-313X.1987.tb00375.x.pd

Normal levels of p27Xic1 are necessary for somite segmentation and determining pronephric organ size

The Xenopus laevis cyclin dependent kinase inhibitor p27Xic1 has been shown to be involved in exit from the cell cycle and differentiation of cells into a quiescent state in the nervous system, muscle tissue, heart and retina. We show that p27Xic1 is expressed in the developing kidney in the nephrostomal regions. Using over-expression and morpholino oligonucleotide (MO) knock-down approaches we show normal levels of p27Xic1 regulate pronephros organ size by regulating cell cycle exit. Knock-down of p27Xic1 expression using a MO prevented myogenesis, as previously reported; an effect that subsequently inhibits pronephrogenesis. Furthermore, we show that normal levels of p27Xic1 are required for somite segmentation also through its cell cycle control function. Finally, we provide evidence to suggest correct paraxial mesoderm segmentation is not necessary for pronephric induction in the intermediate mesoderm. These results indicate novel developmental roles for p27Xic1, and reveal its differentiation function is not universally utilised in all developing tissues

Rhesus TRIM5α disrupts the HIV-1 capsid at the inter-hexamer interfaces

TRIM proteins play important roles in the innate immune defense against retroviral infection, including human immunodeficiency virus type-1 (HIV-1). Rhesus macaque TRIM5α (TRIM5αrh) targets the HIV-1 capsid and blocks infection at an early post-entry stage, prior to reverse transcription. Studies have shown that binding of TRIM5α to the assembled capsid is essential for restriction and requires the coiled-coil and B30.2/SPRY domains, but the molecular mechanism of restriction is not fully understood. In this study, we investigated, by cryoEM combined with mutagenesis and chemical cross-linking, the direct interactions between HIV-1 capsid protein (CA) assemblies and purified TRIM5αrh containing coiled-coil and SPRY domains (CC-SPRYrh). Concentration-dependent binding of CC-SPRYrh to CA assemblies was observed, while under equivalent conditions the human protein did not bind. Importantly, CC-SPRYrh, but not its human counterpart, disrupted CA tubes in a non-random fashion, releasing fragments of protofilaments consisting of CA hexamers without dissociation into monomers. Furthermore, such structural destruction was prevented by inter-hexamer crosslinking using P207C/T216C mutant CA with disulfide bonds at the CTD-CTD trimer interface of capsid assemblies, but not by intra-hexamer crosslinking via A14C/E45C at the NTD-NTD interface. The same disruption effect by TRIM5αrh on the inter-hexamer interfaces also occurred with purified intact HIV-1 cores. These results provide insights concerning how TRIM5α disrupts the virion core and demonstrate that structural damage of the viral capsid by TRIM5α is likely one of the important components of the mechanism of TRIM5α-mediated HIV-1 restriction. © 2011 Zhao et al

Accounting Problems Under the Excess Profits Tax

DNA vaccines based on subunits from pathogens have several advantages over other vaccine strategies. DNA vaccines can easily be modified, they show good safety profiles, are stable and inexpensive to produce, and the immune response can be focused to the antigen of interest. However, the immunogenicity of DNA vaccines which is generally quite low needs to be improved. Electroporation and co-delivery of genetically encoded immune adjuvants are two strategies aiming at increasing the efficacy of DNA vaccines. Here, we have examined whether targeting to antigen-presenting cells (APC) could increase the immune response to surface envelope glycoprotein (Env) gp120 from Human Immunodeficiency Virus type 1 (HIV- 1). To target APC, we utilized a homodimeric vaccine format denoted vaccibody, which enables covalent fusion of gp120 to molecules that can target APC. Two molecules were tested for their efficiency as targeting units: the antibody-derived single chain Fragment variable (scFv) specific for the major histocompatilibility complex (MHC) class II I-E molecules, and the CC chemokine ligand 3 (CCL3). The vaccines were delivered as DNA into muscle of mice with or without electroporation. Targeting of gp120 to MHC class II molecules induced antibodies that neutralized HIV-1 and that persisted for more than a year after one single immunization with electroporation. Targeting by CCL3 significantly increased the number of HIV-1 gp120-reactive CD8(+) T cells compared to non-targeted vaccines and gp120 delivered alone in the absence of electroporation. The data suggest that chemokines are promising molecular adjuvants because small amounts can attract immune cells and promote immune responses without advanced equipment such as electroporation.Funding Agencies|Research Council of Norway; Odd Fellow</p

Ebola Zaire Virus Blocks Type I Interferon Production by Exploiting the Host SUMO Modification Machinery

Ebola Zaire virus is highly pathogenic for humans, with case fatality rates approaching 90% in large outbreaks in Africa. The virus replicates in macrophages and dendritic cells (DCs), suppressing production of type I interferons (IFNs) while inducing the release of large quantities of proinflammatory cytokines. Although the viral VP35 protein has been shown to inhibit IFN responses, the mechanism by which it blocks IFN production has not been fully elucidated. We expressed VP35 from a mouse-adapted variant of Ebola Zaire virus in murine DCs by retroviral gene transfer, and tested for IFN transcription upon Newcastle Disease virus (NDV) infection and toll-like receptor signaling. We found that VP35 inhibited IFN transcription in DCs following these stimuli by disabling the activity of IRF7, a transcription factor required for IFN transcription. By yeast two-hybrid screens and coimmunoprecipitation assays, we found that VP35 interacted with IRF7, Ubc9 and PIAS1. The latter two are the host SUMO E2 enzyme and E3 ligase, respectively. VP35, while not itself a SUMO ligase, increased PIAS1-mediated SUMOylation of IRF7, and repressed Ifn transcription. In contrast, VP35 did not interfere with the activation of NF-κB, which is required for induction of many proinflammatory cytokines. Our findings indicate that Ebola Zaire virus exploits the cellular SUMOylation machinery for its advantage and help to explain how the virus overcomes host innate defenses, causing rapidly overwhelming infection to produce a syndrome resembling fulminant septic shock

T cell–derived inducible nitric oxide synthase switches off TH17 cell differentiation

RORγt is necessary for the generation of TH17 cells but the molecular mechanisms for the regulation of TH17 cells are still not fully understood. We show that activation of CD4+ T cells results in the expression of inducible nitric oxide synthase (iNOS). iNOS-deficient mice displayed enhanced TH17 cell differentiation but without major effects on either TH1 or TH2 cell lineages, whereas endothelial NOS (eNOS) or neuronal NOS (nNOS) mutant mice showed comparable TH17 cell differentiation compared with wild-type control mice. The addition of N6-(1-iminoethyl)-l-lysine dihydrochloride (L-NIL), the iNOS inhibitor, significantly enhanced TH17 cell differentiation, and S-nitroso-N-acetylpenicillamine (SNAP), the NO donor, dose-dependently reduced the percentage of IL-17–producing CD4+ T cells. NO mediates nitration of tyrosine residues in RORγt, leading to the suppression of RORγt-induced IL-17 promoter activation, indicating that NO regulates IL-17 expression at the transcriptional level. Finally, studies of an experimental model of colitis showed that iNOS deficiency results in more severe inflammation with an enhanced TH17 phenotype. These results suggest that NO derived from iNOS in activated T cells plays a negative role in the regulation of TH17 cell differentiation and highlight the importance of intrinsic programs for the control of TH17 immune responses