Hapten-specific responses to the phenyltrimethylamino hapten. III. Mice whose delayed-type hypersensitivity responses cannot be abrogated by the presence of anti-idiotypic suppressor T cells lack a critical modulatory T cell function.

A single intraperitoneal injection of the monovalent synthetic antigen, tyrosinated trimethylaminoaniline [tyr(TMA)] in Freund's complete adjuvant induces an antiidiotypic second-order T suppressor (Ts2) cell population 6 wk later. This population was able to suppress TMA-specific delayed-type hypersensitivity (DTH) responses when adoptively transferred into normal syngeneic recipients. However, they failed to function intrinsically. The inability of the Ts2 to function intrinsically was not caused by compensating idiotype-negative T cells that mediate DTH. Rather, this paradoxical observation was found to be caused by the absence or loss of function of a critical modulatory T cell population in the suppressor cell-bearing mice. This cell is functionally active in normal mice immunized for DTH responses and is sensitive to cyclophosphamide treatment. In addition, this cell type bears idiotype on its surface and is Thy-1+ and Lyt-1-,2+. It was demonstrated that by adoptively transferring the activated modulatory T cells from normal mice into tyr(TMA)-immune recipients, it was possible to observe suppressor cell function intrinsically. The potential importance of modulatory T cell function in the regulation of antibody and DTH responses is discussed.

T cell replacing factor substitutes for an I-J+ idiotype-specific T helper cell.

An in vitro system for the study of idiotype (Id) expression on antitrimethylamino hapten antibody-producing cells and its regulation by two classes of helper T cells is described. These cells are distinguished in four ways: one requires a hapten-carrier bridge and gives a good response that is low in Id; it does not bind to Id-coated dishes and is not affected by anti-I-J plus complement. The other requires antigen but not a hapten-carrier bridge, is bound by Id-coated dishes and is killed by anti-I-J and complement. The Id-specific cell appears to be antigen specific and acts via a soluble factor(s).

Anti-phenyltrimethylamino immunity in mice. II. L-Tyrosine-p-azophenyltrimethylammonium-induced suppressor T cells selectively inhibit the expression of B-cell clones bearing a cross-reactive idiotype.

The anti-phenyltrimethylamino (TMA) response in A/J mice is characterized by a cross-reactive idiotype(s) (CRI) that appears linked to the Ig-Ie allotype. These findings made it attractive to look for a CRI on T cells reactive to the same TMA determinant. Thus a suppressor T-cell (Ts) assay specific for L-tyrosine-p-azophenyltrimethylammonium [tyr(TMA)] was developed. A/J mice were primed with either tyr(TMA) in complete Freund's adjuvant (CFA), L-tyrosine-azobenzenearsonate [tyr(ABA)] in CFA, or with CFA alone. 6 wk later all mice were inoculated with TMA-bovine serum albumin (BSA) in CFA, boosted with soluble TMA-BSA 3 wk later, and plaqued 7 d after the soluble boost. Priming with tyr(TMA) in CFA resulted in 66% suppression of anti-TMA plaque-forming cells (PFC) as compared with control groups primed with tyr(ABA) in CFA or CFA alone. The suppression was shown to be mediated by Ts, as only T cells but not B cells from suppressed animals transfer the suppression in adoptive cell transfer experiments into lethally irradiated recipients. The profile of the anti-TMA PFC in the suppressed and nonsuppressed animals was examined via incorporation of anti-idiotypic sera (specific for CRI-TMA) into the plaquing medium. The results of these experiments indicate that the suppression of the major CRI+-TMA PFC was virtually complete, whereas the CRI--TMA PFC are left intact. When A/J mice were primed with idiotypic antisera (anti-Id) or normal rabbit serum (NRS) rather than with the antigen on CFA alone, and the same protocol was followed thereafter, the anti-Id-inoculated mice were suppressed by 63% when compared with the NRS-primed controls. Again the suppression could be accounted for by the exclusive elimination of CRI+ anti-TMA PFC. The possibility that the antigen-induced idiotype suppression may result from idiotypic restrictions between interacting CRI+-Ts and CRT+-B cells will be discussed.

Inhibition of T-antigen-binding cells by idiotypic antisera.

Shared idiotypy between B- and T-cell receptors specific for the antigen L-tyrosine-p-azophenyltrimethylammonium [tyr(TMA)] was studied in an antigen-binding assay using idiotypic antisera. These idiotypic reagents were prepared by inoculation of rabbits with purified anti-tyr(TMA) antibody raised in strain 13 guinea pigs. The antisera blocked 78-83% of the antigen-binding T cells (T-ABC) and 50-55% of the antigen-binding B cells (B-ABC) from tyr(TMA)-immune strain 13 and outbred lymph node cells (LNC). An excess of normal guinea pig Ig in the ABC assay did not affect the ability of the idiotypic antisera to block T- and B-ABC. Nylon wool-passed tyr(TMA)-immune LNC were trypsin treated resulting in a 75% loss of T-ABC. The trypsin-treated population was then cultured for 16 h which resulted in a return of T-ABC to 92% of pretrypsin values. 77% of these regenerated T-ABC could be blocked with idiotypic antisera. Specificity of the idiotypic antisera was tested in L-tyrosine-p-azobenzenearsonate-immune guinea pig LNC. Neither T- nor B-ABC were blocked in this heterologous system. Further blocking experiments were performed to characterize the nature of the T-ABC receptor. A variety of anti-Ig reagents, some of which block B-ABC, do not inhibit T-ABC suggesting that variable regions on T cells are not linked to Ig Constant regions.

T cell subsets, epitope mapping, and HLA-restriction in patients with allergic bronchopulmonary aspergillosis.

Allergic bronchopulmonary aspergillosis (ABPA) is a hypersensitivity lung disease characterized by Aspergillus fumigatus (Af) colonization, IgE and IgG anti-Af antibodies, pulmonary infiltrates, bronchiectasis, and pulmonary fibrosis. Little is known regarding T cell responses and their role in the pathogenesis of ABPA. To examine T cell reactivity to Af antigens, T cell clones (TCC) specific to the Asp f 1 antigen, an 18-kD protein of Af, were established from the peripheral blood of three ABPA patients. The majority of TCC isolated from ABPA patients, and specific for the Asp f 1 allergen of Af, are IL-4 producing CD4+ cells of the Th2 phenotype. Further analysis in this study revealed that the majority of TCC reacted to mainly two epitopes of Asp f 1, while the remaining TCC reacted to three additional "minor" epitopes. Blocking studies using monoclonal antibodies specific for class II HLA-D region gene products showed that most TCC, 19/21, were restricted by HLA-DR molecules, and the remaining two clones by HLA-DP molecules. The use of a panel of HLA-matched and mismatched EBV-transformed B cells as antigen presenting cells revealed that the HLA-DR restriction was mediated exclusively by either the HLA-DR2 or HLA-DR5 alleles. Genotyping of DRB1 gene products showed that class II presentation for most clones was not restricted to a single allele, representing DRB1 gene products of either HLA-DR2 or DR5. These studies offer insight into the cellular and molecular determinants which contribute to the immunopathophysiology of ABPA.

A novel cis-acting element required for lipopolysaccharide-induced transcription of the murine interleukin-1 beta gene.

Regulatory elements important for transcription of the murine interleukin-1 beta (IL-1 beta) gene lie within a DNase I-hypersensitive region located > 2,000 bp upstream from the transcription start site. We have identified within this region a novel positive regulatory element that is required for activation of an IL-1 beta promoter-chloramphenicol acetyltransferase (CAT) fusion gene in the murine macrophage line RAW264.7. Electrophoretic mobility shift analysis of the 3' portion (-2315 to -2106) of the hypersensitive region revealed at least two nuclear factor binding sites, one of which is located between positions -2285 and -2256. Competitive inhibition studies localized the binding site to a 15-bp sequence between -2285 and -2271. Nuclear factor binding was lost by mutation of the 6-bp sequence from -2280 to -2275. The specific retarded complex formed with RAW264.7 nuclear extract was not detected under similar conditions with nuclear extracts from RLM-11, a murine T-cell line which does not express IL-1 beta RNA. Mutation of the 6-bp sequence (-2280 to -2275) in the chimeric IL-1 beta promoter -4093 +I CAT plasmid virtually eliminated the activation of this reporter gene by lipopolysaccharide (LPS) in transfected RAW264.7 cells. Multimerization of the 15-bp sequence containing the core wild-type 6-bp sequence 5' of minimal homologous or heterologous promoters in CAT reporter plasmids resulted in significant enhancement of CAT expression compared with parallel constructs containing the mutant 6-bp core sequence. This element was LPS independent and position and orientation dependent. The multimerized 15-bp sequence did not enhance expression in RLM-11 cells. Methylation interference revealed contact residues from -2281 to -2271, CCAAAAAGGAA. Because a search of the NIH TFD data bank with the 11-bp binding site sequence found no homology to known nuclear factor binding sites, we have designated this sequence the IL1 beta -upstream nuclear factor 1 (IL1 beta -UNF1) target. UV cross-linking and sodium dodecyl sulfate-polyacrylamide electrophoresis identified an IL1 beta -UNF1-specific binding factor approximately 85 to 90 kDa in size.

In vivo host immune to a tumor-specific transplantation antigen induced by Rous sarcoma virus.

We examined the host immune response to a tumor-specific transplantation antigen (TSTA) induced by Rous sarcoma virus (RSV) In vivo. In contrast to previous in vitro studies, the present investigation demonstrated in vivo host immunity of the TSTA 10-55 days after tumor inoculation. Immunity to the TSTA appeared specific, since the homologous RSV tumor was rejected. whereas the heterologous tumor grew progressively. No generalized suppression of cell-mediated or hymoral immunity was shown, because tumor-bearing hosts retained the ability to reject heterologous tumor cells and mounted a normal plaque response to sheep red blood cells. Although alpha-globulin levels were elevated, they did not appear to affect the host's immunity to the growing tumor or to heterologous antigens. Assoicated with the progressively growing tumor was the appearance in the serum of a fetal antigen with characteristics of an alpha-2 acute phase protein.

Glucocorticoid effects on lipopolysaccharide-stimulated murine B-cell leukemia line (BCL1) cells.

The effects of glucocorticoids were studied in lipopolysaccharide (LPS)-stimulated splenic murine B-cell leukemia line one (BCL1) cells. At 24 hr, LPS caused a 3-fold increase in [3H]-thymidine incorporation compared to similarly cultured unstimulated cells. Triamcinolone acetonide (TA) and dexamethasone at a concentration of 10(-8) M reduced [3H]thymidine incorporation 80 and 53%, respectively, while estradiol at concentrations of 10(-10) to 10(-5) M had no effect. A 500-fold excess of cortexolone or progesterone blocked the response of 10(-8) M TA by 42 and 38%, respectively, indicating that the glucocorticoid response could be inhibited by antiglucocorticoids. The maximum rate of thymidine incorporation in LPS-stimulated cells occurred at 24 hr, a time at which 10(-5) M TA present in parallel cultures from the initiation of LPS stimulation showed a 79% reduction in [3H]thymidine incorporation. If TA was added at any time after the initiation of LPS stimulation, the degree of decrease in nucleotide incorporation was not as marked. Therefore, maximum TA effect in LPS-stimulated BCL1 cells occurred when TA was added to cultures at the onset of mitogen stimulation. We also measured glucocorticoid-specific receptor in whole cells both before LPS stimulation and in BCL1 cells incubated 24 hr in the presence of LPS. The equilibrium dissociation constant, the number of sites/cell, and the hormone specificity of the glucocorticoid receptor were similar prior to and at the peak of mitogen stimulation.

Binding of [3H]triamcinolone acetonide-receptor complexes to chromatin from the B-cell leukemia line, BCL1.

The binding characteristics of partially purified glucocorticoid receptor complexes from hormone sensitive, non-differentiating BCL1 cells to sequentially deproteinized BCL1 chromatin-cellulose was investigated. [3H]Triamcinolone acetonide (TA)-receptor complexes were purified (approx. 30-fold) from DEAE-cellulose columns by salt elution which allowed receptor activation only in the absence of molybdate. Addition of 10 mM molybdate completely blocked salt activation. The binding pattern of the activated [3H]TA-receptor complexes to chromatin-cellulose extracted with 0-8 M guanidine hydrochloride revealed three regions of increased binding activity (acceptor sites), at 2, 5 and 7 M guanidine hydrochloride. Acceptor site binding was markedly reduced for chromatin extracted with 3, 6 and 8 M guanidine hydrochloride. Non-activated receptor complexes demonstrated very low binding to deproteinized chromatin. It was also shown that chromatin binding required glucocorticoid receptors and that free ligand or ligand bound to other proteins did not bind significantly to chromatin. In addition, binding of [3H]TA-receptor complexes to partially deproteinized chromatin was competable by unlabeled TA-receptor complexes. Scatchard analysis demonstrated that chromatin from non-differentiating BCL1 cells possesses multiple, high-affinity binding sites which differ in their affinity for the glucocorticoid receptor. Partially deproteinized chromatin from lipopolysaccharide-stimulated BCL1 cells demonstrated a different pattern of receptor binding, i.e., receptor binding was significantly greater to chromatin previously extracted with 6-8 M guanidine hydrochloride. These results suggest that differentiation alters the state of chromatin and the interaction of non-histone protein/DNA acceptor sites with glucocorticoid receptors. These alterations may play a role in the acquisition of hormone resistance.

Effect of estrogens on IL-1beta promoter activity.

It is well documented that steroid hormones modulate cytokine gene expression. In some tissues estrogens are known to suppress cytokine production while in other tissue types, cytokine expression is enhanced by the hormone. This study was conducted to investigate the regulatory mechanisms which underlie the modulation of the interleukin-1beta (IL-1beta) gene at the transcription level. To accomplish this, the macrophage cell line RAW264.7, which appeared insensitive to 17beta-estradiol (E2) treatment, was stably transfected with the human estrogen receptor (ER) and an IL-1beta promoter-CAT reporter construct. E2 markedly enhanced LPS-induced IL-1beta promoter-driven CAT activity in an E2 dose dependent manner. This responsiveness was estrogen specific since no synergism was observed between LPS and the sex steroids testosterone or progesterone while the estrogen analogue 17alpha-estradiol stimulated only at 10 to 100 times the amount required for 17beta-E2. Several antiestrogens, H1285, ICI 182 780, and tamoxifen inhibited the estrogen stimulated enhancement of IL-1beta promoter activity in a dose-dependent manner, indicating that this effect was indeed mediated through the ER in a ligand dependent manner. The estrogenic effect appeared to be indirect and time dependent since the addition of E2 was required hours prior to LPS stimulation; addition of E2 and LPS at the same time resulted in a greatly reduced estrogenic effect. The estrogen metabolites 17-epiestriol and 16-keto-17beta-E2 displayed an estrogenic response virtually indistinguishable from E2. 4-Hydroxyestradiol displayed activity only at 100-fold the concentration of E2 while 2-hydroxyestrone showed no activity at any of the concentrations tested. Overall the results demonstrate that E2 and some metabolites of E2 synergize with LPS to markedly enhance IL-1beta promoter activity through ER mediated processes.

An NFIL-6 sequence near the transcriptional initiation site is necessary for the lipopolysaccharide induction of murine interleukin-1 beta.

We have analyzed the interleukin-1 beta (IL-1 beta) promoter region near the cap site. Specific DNA sequences required for lipopolysaccharide (LPS) induction within this region were identified using transfection of reporter plasmids that contained portions of the proximal IL-1 beta 5'-flanking sequence. An LPS-responsive activation area was localized between nucleotides -50 to -100, and down-regulating sequences were present between nucleotides -100 and -2,111. A NFIL-6 site between -92 and -84 was identified in the functionally active region. Base substitutions within this single NFIL-6 site in the context of a 4.1-kb IL-1 beta promoter segment resulted in dramatic reduction of LPS-induced gene transcription. Introduction of multimers of this NFIL-6 sequence immediately 5' to minimal homologous or heterologous promoters conferred LPS inducibility in each case. Anti-C/EBP beta (NFIL-6) and anti-C/EBP delta (NFIL-6 beta) antibodies identified both of these proteins in complexes formed between the NFIL-6 site and mononuclear cell nuclear extracts. These data show that the proximal NFIL-6 site is required for the activation of murine IL-1 beta gene expression by endotoxin.

Depletion of murine anti-azobenzenearsonate plaque-forming cells by derivatized polyacrylamide beads.

The depletion of secondary p-azobenzenearsonate plaque-forming cells (ABA-PFC) by affinity columns substituted with ABA was dependent on the primary dose, times elapsed after priming, and the nature of the side-arm on the columns. Thus, higher priming doses of ABA coupled to keyhole limpet haemocyanin (50-500 micrograms of ABA-KLH in complete Freund's adjuvant (CFA] favoured depletion of ABA-PFC by ABA coupled to a 6- aminocaproyltyrosine side-arm (SAC-TYR-ABA beads), whereas ABA-PFC from mice primed with 1 microgram of ABA-KLH in CFA were never depleted; this latter population were only depleted by ABA coupled to a histamine side-arm (HIST-ABA beads) 7 months after priming. These data are consistent with the notion that two specificities, HIST-ABA and TYR-ABA, appear to emerge independently, lower priming doses inducing the preferential appearance of the HIST-ABA specificity.

Extensive cross-reactive idiotypy among guinea-pig anti-TMA antibodies.

The extensive cross-reactivity of the immune response by strain 13 (S-13) and outbred guinea-pigs (GP) to either the bivalent antigen L-tyrosine-p-azophenyltrimethylammonium (tyr(TMA)) [H-L-tyr(TMA)-NH-(CH2)3]2 (T-S-T) or the conjugate 3-(p-trimethylphenylazo)-N-acetyl-L-tyrosylglycylglycine-bovine serum albumin (T-S-B) was examined. Antibody (Ab) isolated from a single T-S-B immune S-13 animal by affinity chromatography (proband) was used to induce anti-sera (anti-Id) in a rabbit and a syngeneic S-13 GP. These anti-Ids were both shown to be idiotype specific by means of a tube binding radioimmunoassay (TBRIA). A series of S-13 anti-(T-S-B), S-13 anti-(T-S-T), and outbred anti-(T-S-B) antisera were tested as inhibitors in the TBRIA through a range of 6-200 ng of anti-TMA Ab equivalent. Each was found to inhibit extensively, indicating a considerable amount of cross-reactivity of idiotypes present in the sera. In general, the rabbit anti-Id recognized more cross-reactivity among the various antisera than did the GP reagent, particularly among those from T-S-B immune S-13 animals. Addition of free "hapten" tyr(TMA) as an inhibitor specifically displaced 55% of the label from the rabbit reagent at 10(-5)M, while only 25% of binding of the proband to GP anti-Id was inhibitable at this concentration. These findings suggest that the rabbit and GP anti-Ids are directed predominantly to different determinants, the rabbit recognizing the less variant, hapten-binding portion of the GP Ab more than does the syngeneic GP reagent. Data from isoelectric focus patterns support the impression of a highly restricted Ab response to TMA. While each serum gives a distinct binding pattern, several bands appear shared by all responders.

Hapten-specific responses to the phenyltrimethylamino hapten. IV. Occurrence of mechanistically distinct idiotypic suppressor T cells before the appearance of anti-idiotypic suppressor T cells induced by the monovalent antigen L-tyrosine-p-azophenyltrimethylammonium.

We have previously shown that a single i.p. injection of the monovalent synthetic antigen, L-tyrosine-p-azophenyltrimethylammonium [tyr(TMA)] in complete Freund's adjuvant induces an anti-idiotypic T suppressor cell (Ts2) population that can be detected 6 wk later by its ability to shut down delayed-type hypersensitivity (DTH) specific for the TMA hapten. In this paper we present evidence that 2 wk after tyr(TMA) administration, a subset of Ts, termed Ts1, appears that is both functionally and phenotypically distinct from the late appearing Ts2 population. The early occurring Ts1 act only at the induction phase of the DTH response and can also suppress this response intrinsically. This latter point is in marked contrast to our previous observation that the tyr(TMA)-induced anti-idiotypic Ts2 fail to function intrinsically and can only be detected upon adoptive transfer into naive mice. Ts1 bear idiotypic receptors and are Ly-1+,2- in contrast to the anti-idiotypic Ly-1-,2+ Ts2 population. In addition, unlike the Ts2 population, Ts1 are comparatively nylon wool-adherent. Adsorption of Ts1 on either antigen- or idiotype-coated petri dishes indicate that the suppressor activity can be transferred only by antigen-binding cells. Cellfree factors prepared from spleens containing the Ts1 population can suppress DTH only if administered at the induction phase of the response, in contrast to the factors derived from the Ts2 population that act both at induction as well as effector phases, suggesting that Ts1 and Ts2 can function via soluble mediators. Finally, we show that when Ts1-bearing mice are primed and boosted for anti-TMA antibody formation, the resulting response was overall reduced with respect to the idiotype-positive and negative plaque-forming cells that differs from the Ts2-bearing hosts wherein the idiotypic component is preferentially suppressed. The appearance of Ts1 before the detection of Ts2 in the same experimental animals is discussed with reference to a normal physiologic sequence of events involved in suppressor pathways.

Hapten-specific responses to the phenyltrimethyl-amino hapten. I. Evidence for idiotype-anti-idiotype interactions in delayed-type hypersensitivity in mice.

The delayed-type hypersensitivity (DTH) reaction to the phenyltrimethylamino (TMA) hapten in mice has been investigated. TMA-derivatized syngeneic spleen cells (TMA-SC) administered s.c. in several strains of mice consistently evoked DTH reactivity, as measured by footpad swelling after challenge with the diazonium salt of TMA. DTH could be induced by low levels of anti-idiotypic antisera (anti-Id) in lieu of antigen. The DTH reaction induced by either mode was hapten-specific, could be transferred into naive recipients by viable lymph node cells but not with serum from immune mice and was not influenced by cyclophoshamide pretreatment. Unlike TMA-SC which induced DTH in all of the strains of the mice tested, anti-Id induced DTH only in strains of the Igh-1e allotype. Positive DTH reactions were induced by anti-Id in the C57.Ige strain (H-2b, Igh-1e) but not in its allotype-congenic partner C57BL/6J (H-2b, Igh-1b). Interestingly this reaction could be suppressed if relatively high amounts of anti-Id were inoculated i.v. just prior to antigen challenge. In addition, the administration of anti-Id 1 h prior to antigen challenge in TMA-SC-sensitized mice significantly blocked the DTH reaction only in the Igh-1e strains. These results demonstrate that the induction and abrogation of TMA-specific DTH by anti-Id is linked to the IgCh locus.

Hapten-specific responses to the phenyltrimethylamino hapten. V. A single chain antigen-binding I-J+ first-order T suppressor factor requires antigen to induce anti-idiotypic second-order suppressor T cells.

We have previously shown that a single i.p. injection of the monovalent antigen, L-tyrosine-p-azophenyltrimethylammonium in complete Freund's adjuvant induces a Ly-1+2-, idiotype-bearing, and antigen-binding first-order T suppressor (Ts1) population. We showed that soluble factors extracted from these cells could suppress delayed-type hypersensitivity responses if administered at the induction phase of the response. In this paper we additionally characterize the suppressor factor, TsF1, with respect to its biologic, serologic, and chemical properties. The studies show that the TsF1 is neither allotype nor H-2 restricted and can induce anti-idiotypic T suppressor cells (Ts2), but it requires the presence of antigen to do so. The factor binds antigen, bears I-J encoded determinants, is resistant to reduction and alkylation, and elutes as a single chain factor after adsorption onto monoclonal anti-I-J antibody-coupled Sepharose beads in the presence of dithiothreitol (DTT). This is in marked contrast to TsF2 (derived from Id-specific Ts2-containing spleen cells), which lost its suppressive activity after reduction and alkylation, and behaves as a two chain factor after adsorption and elution from anti-I-J-coupled beads in the presence of DTT. The TsF1 is discussed with respect to the properties of it and those of TsF1 from other similar idiotype-dominated antigen systems.

Idiotype-specific T suppressor factor alternatively interacts with a nonspecific T-acceptor-like cell to mediate immune suppression.

The ability of the idiotype (Id)-specific second-order T suppressor factor (TsF2) to interact with a final effector Ts cell type other than the previously reported third-order Ts (Ts3) subset was studied in the phenyltrimethylamino (TMA) hapten system. Hence, mice were primed with unrelated heterologous haptens to induce the nonspecific T acceptor (Tacc) cells following published procedures. When enriched T cell populations containing these nonspecific Ts were briefly incubated in vitro with TMA-TsF2, they produced suppression upon adoptive transfer into cyclophosphamide-treated mice which had been previously immunized for TMA-specific delayed-type hypersensitivity. Despite the fact that the effector population studied in this report also required Id-binding TsF2 for its function, it differs markedly from the Ts3 subset studied previously in the TMA system. First, the cell type studied herein could be easily generated with noncrossreacting heterologous chemically reactive haptens when applied directly to the skin of mice. Furthermore, these Ts effector cells had no detectable intrinsic receptors for homologous haptens and most importantly, unlike Ts3, this population had no affinity for the TMA hapten. Nevertheless, the nonspecifically induced Ts once activated by TsF2 suppresses TMA-directed, but not similar immune responses specific for heterologous haptens. Thus the results indicate that TsF2 can functionally interact with a final effector Ts subset (very similar to the Tacc) other than the well described Ts3 population. The ramifications of these findings are discussed with reference to a generalized view of the cellular basis of terminal phases of immune suppression.