Soluble factors in tolerance and contact sensitivity to DNFB in mice. X. IL-2 is the activation signal mediating release of synthesized suppressor factor

Two signals are required for the in vitro activation of Lyt2+ T suppressor cells (Ts) from mice tolerized with 2,4-dinitrobenzene sulfonate (DNBS) to produce soluble suppressor factors (SSF) which suppress the transfer of contact sensitivity to dinitrofluorobenzene (DNFB). Recognition of DNP/class I MHC (signal one) stimulates the Ts to synthesize SSF. Release of SSF requires a soluble mediator (signal two) produced by the interaction of L3T4+ T cells from tolerant mice with I-A on metabolically functional cells in the DNP-presenting cell population. The purpose of this study was to examine the nature of this second Ts activation signal. Coculture of tolerant spleen cells and glutaraldehyde-fixed (Glu-) DNP-labeled spleen cells (DNP-SC) resulted in the synthesis but not release of SSF. Addition of either IL-1 or IL-2 to these cultures induced SSF release. Treatment of such cultured cells with the anti-murine IL-2 receptor antibody PC 61.5.3 blocked the IL-2- and IL-1-stimulated release of SSF. Release of SSF was also blocked when tolerant cells were cultured with (unfixed) DNP-SC in the presence of a monoclonal anti-IL-2 antibody. IL-2 but not IL-1 was able to stimulate the Ts to release synthesized SSF in the absence of L3T4+ TH activity. First, addition of IL-2 to cocultures of tolerant cells and DNP-presenting I-A- cells induced release of the synthesized SSF, whereas addition of IL-1 did not. Second, IL-2 also stimulated SSF release in cocultures of L3T4+ T cell-depleted tolerant cells and Glu-DNP-SC, whereas IL-1 did not. Tolerant cells pretreated with IL-2 and then washed were able to synthesize and release SSF upon culture with Glu-DNP-SC. Pretreatment of tolerant cells with IL-1 did not stimulate SSF release upon subsequent culture with Glu-DNP-SC. These results indicate that the Lyt2+ Ts from DNBS-tolerant mice express IL-2 receptors and IL-2 is the lymphokine which induces the Ts to release synthesized SSF. Thus, IL-2 provides a differentiative signal during the functional activation of these regulatory T cells.     

Trade-offs in carbon-degrading enzyme activities limit long-term soil carbon sequestration with biochar addition

Biochar amendment is one of the most promising agricultural approaches to tackle climate change by enhancing soil carbon (C) sequestration. Microbial-mediated decomposition processes are fundamental for the fate and persistence of sequestered C in soil, but the underlying mechanisms are uncertain. Here, we synthesise 923 observations regarding the effects of biochar addition (over periods ranging from several weeks to several years) on soil C-degrading enzyme activities from 130 articles across five continents worldwide. Our results showed that biochar addition increased soil ligninase activity targeting complex phenolic macromolecules by 7.1%, but suppressed cellulase activity degrading simpler polysaccharides by 8.3%. These shifts in enzyme activities explained the most variation of changes in soil C sequestration across a wide range of climatic, edaphic and experimental conditions, with biochar-induced shift in ligninase:cellulase ratio correlating negatively with soil C sequestration. Specifically, short-term (<1 year) biochar addition significantly reduced cellulase activity by 4.6% and enhanced soil organic C sequestration by 87.5%, whereas no significant responses were observed for ligninase activity and ligninase:cellulase ratio. However, long-term (≥1 year) biochar addition significantly enhanced ligninase activity by 5.2% and ligninase:cellulase ratio by 36.1%, leading to a smaller increase in soil organic C sequestration (25.1%). These results suggest that shifts in enzyme activities increased ligninase:cellulase ratio with time after biochar addition, limiting long-term soil C sequestration with biochar addition. Our work provides novel evidence to explain the diminished soil C sequestration with long-term biochar addition and suggests that earlier studies may have overestimated soil C sequestration with biochar addition by failing to consider the physiological acclimation of soil microorganisms over time.     

Soluble factors in tolerance and contact sensitivity to 2,4-dinitro-fluorobenzene in mice. IX. A monoclonal T cell suppressor molecule is structurally and serologically related to the alpha/beta T cell receptor

Ts cells from mice tolerized with dinitrobenzene sulfonate produce a DNP-specific, MHC-restricted soluble suppressor factor (SSF) which regulates contact sensitivity to 2,4-dinitro-fluorobenzene. Previous studies have shown that the SSF-producing T cells and the soluble factor have the same hapten/MHC specificity suggesting that SSF may represent a secreted form of the Ts membrane receptor. The relationship between TCR proteins and SSF was investigated by examining the structural and serologic properties of a monoclonal DNP/H-2Kd-specific suppressor molecule produced by a Ts hybridoma. Reduction followed by alkylation abrogated the ability of the 3-10 molecule to inhibit transfer of contact sensitivity to 2,4-dinitro-fluorobenzene, indicating that intact disulfide bonds were a required structural property for suppression. Reduction of the 3-10 molecule followed by affinity chromatography on DNP-coupled Sepharose beads indicated that the 3-10 suppressor molecule is a dimer and that one of its chains binds to cell-free DNP. Serologic properties of the 3-10 molecule were examined by determining the ability of pan-reactive rabbit anti-TCR antibodies and anti-V beta 8 mAb KJ16.133 and F23.1 to adsorb suppressor activity from 3-10 culture supernatant and affinity purified 3-10 ascites material. All three reagents adsorbed the suppressor activity whereas control antibodies had no effect. When 3-10 material was passed through a F23.1-conjugated Sepharose affinity column, suppressor activity was recovered in the column eluate but not in the effluent fraction. When the 3-10 molecule was reduced and separated into its two chains (i.e., DNP-binding and non-DNP-binding chains), it was found that the anti-V beta 8 antibody F23.1-bound to the non-DNP-binding chain of the suppressor molecule. Collectively, these results indicate that the monoclonal 3-10 suppressor molecule is structurally similar to the alpha/beta TCR and suggest that the 3-10 molecule expresses a determinant encoded by the V beta 8 family of TCR genes. These results are consistent with our hypothesis that these suppressor molecules represent a secreted form of the TCR expressed on the surface of the DNP-specific Ts.     

Tolerance and contact sensitivity to DNFB in mice. VII. Functional demonstration of cell-associated tolerogen in lymph node cell populations containing specific suppressor cells.

Adoptive tolerance to contact sensitivity to DNFB is mediated by suppressor T cells. These cells are induced by iv injection of the hapten DNB-SO₃. Experiments were carried out to investigate the question of simultaneous transfer of tolerogen (DNB-SO₃ or its conjugation product DNP) with the suppressor cells. The results showed that tolerant lymph node cells pretreated in vitro with anti-TNP serum before transfer were unable to induce unresponsiveness to DNFB. Tolerant cells treated with either anti-TNP serum which had been passed over a TNP-affinity column or with polyvalent anti-immunoglobul in serum were not inhibited. These results functionally demonstrate that LN cell populations containing DNFB suppressor cells have accessible hapten (e.g., DNP) associated with their membrane, which is necessary for induction of adoptive tolerance. The hapten (tolerogen) appears to be bound directly to the cell surface rather than as an immune complex.     

The Role of the <Emphasis Type="Italic">sigB</Emphasis> Gene in the General Stress Response of <Emphasis Type="Italic">Listeria monocytogenes</Emphasis> Varies between a Strain of Serotype 1/2a and a Strain of Serotype 4c

The ability of Listeria monocytogenes to resist many adverse environmental conditions has been attributed in part to activation of the alternative sigma factor ς^B, encoded by the sigB gene. The ability of this pathogen to survive and grow under stress conditions varies between strains within the species. The current study was undertaken to determine whether the role played by the sigB gene in the stress response varies among strains of different serotypes. Null mutations were generated in the sigB genes of L. monocytogenes L61 (serotype 1/2a) and L99 (serotype 4c), and the survival of the resulting mutants was compared with that of the wild-type strains under osmotic, oxidative, and carbon starvation stress conditions and on exposure to bacteriocins, ethanol, acid, and heat. Except in a few cases, strain L61 displayed greater dependence on the sigB products for survival of adverse conditions than did strain L99. The results of this study indicated that the relative importance of the sigB gene in the stress response is not the same in all strains of L. monocytogenes, and this difference may be specific to serotype groupings within the species. Received: 8 May 2002 / Accepted: 27 August 2002     

Antibody-mediated regulation of T cell responses. I. Characterization of a monoclonal antibody which specifically regulates contact hypersensitivity to DNFB in BALB/c mice

Contact hypersensitivity (CS) to 2,4-dinitrofluorobenzene (DNFB) in BALB/c mice is regulated by autoanti-idiotypic antibody. This report describes the preparation and characterization of a monoclonal antibody, 2-16.1, which has characteristics previously described for the serum anti-idiotypic antibodies. Monoclonal 2-16.1 was prepared by fusing lymph node (LN) cells from optimally sensitized BALB/c mice to the P3X myeloma. The monoclonal product of the cloned hybridoma is an IgM (K) immunoglobulin which does not bind to DNP-protein but which does bind to other immunoglobulins with anti-DNP specificity, primarily of the IgM class. Functionally, 2-16.1 inhibits the efferent limb of the CS reaction as measured by passive transfer of immunity. This inhibition is antigen-specific and appears to require the presence of a subset of Ia+ T cells in the DNFB-immune LN cell population. Suppression of transfer of immunity is strain-specific. Finally, suppression occurs only in the absence of complement, indicating that a lytic mechanism is not involved and that 2-16.1 does not recognize determinants expressed on the effector T cells of the CS reaction. Collectively, these results indicate that 2-16.1 is a monoclonal anti-idiotypic antibody, and that the hybridoma CSDNP 2-16.1 represents a clone of B cells which is stimulated during the primary CS response to DNFB and whose antibody product is involved in the endogenous, active regulation of this T cell-mediated response.