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S K Pierce

Publications and source records attributed to S K Pierce.

At least 91 records · Page 5Linked to original sources
PubMed1988

Enhanced T cell responses to antigenic peptides targeted to B cell surface Ig, Ia, or class I molecules.

The helper T cell recognition of soluble globular protein antigens requires that the proteins be processed by an APC, releasing a peptide that is transported to and held on the APC surface where it is recognized by the specific T cell in conjunction with Ia. When cellular processing functions are blocked, APC lose their ability to present native antigens while retaining the capacity to activate T cells when provided with a cognate peptide fragment that contains the T cell antigenic determinant. In this report, we show that a peptide fragment of the soluble globular protein antigen tobacco hornworm moth cytochrome c, residues 92-103 containing an additional NH2-terminal cysteine residue (THMcCys92-103), is effectively presented by B cells to an I-Ek-restricted, THMc-specific T cell hybrid when covalently coupled to antibodies specific for B cell surface Ig, Ia (Ak), or class I (Kk). Maximal activation of the T cells to the THMcCys92-103-antibody conjugates is achieved with 1/100-1/1,000th of the peptide required using unconjugated THMcCys92-103 or THMcCys92-103 coupled to nonspecific antibody. The T cell response to the peptide antibody conjugates is MHC restricted, but unlike native cytochrome c-antibody conjugates, THMcCys92-103-antibody conjugates do not require processing and can be presented by paraformaldehyde-fixed B cells. The THMcCys92-103-antibody conjugate are nearly as effective when incubated with B cells, and the unbound conjugates washed away before addition of T cells as when continuously present in culture with T cells and B cells, indicating that the active peptide antibody conjugate is associated at the B cell surface. The presentation of THMcCys92-103 coupled to monovalent Fab fragments of rabbit anti-Ig antibodies is less effective than that of the peptide coupled to bivalent antibody when either live or fixed B cells are APC, indicating that the avidity for the APC surface afforded by bivalent binding may be important in the conjugate's antigenicity. The results presented here indicate that a T cell-antigenic peptide, covalently coupled to a larger antibody molecule, can be readily recognized by an Ia-restricted helper T cell in the absence of processing. Moreover, the ability of the peptide to bind to B cell surfaces greatly augments the peptide's antigenicity, even when the binding is to structures distinct from the Ia molecule required for T cell activation.

Animals↗
PubMed1988

Time dependence of B cell processing and presentation of peptide and native protein antigens.

Th cell recognition of globular proteins requires the uptake and intracellular processing of the native Ag by an APC to produce a peptide fragment containing the T cell antigenic determinant, which is recognized in conjunction with Ia. This report describes the time course of the processing and presentation of a soluble globular protein Ag, pigeon cytochrome c (Pc), and of the presentation of a C-terminal peptide fragment of Pc, residues 81 to 104 (Pc 81-104), which does not require processing. Splenic B cells, acting as APC, require 6 to 8 h incubation with native Pc to process and present it to an I-Ek-restricted Pc-specific T cell hybrid, resulting in the secretion of IL-2. Moreover, the time required for B cells to process Pc is the same whether the Ag is taken up by nonspecific fluid phase pinocytosis or by binding to surface Ig. Once processed, Ag is lost from the B cell surface by 8 to 12 h, although when provided with fresh Pc, the same B cells are still capable of processing and presenting. In contrast to native Pc, only 1 to 2 h are required for the peptide fragment Pc 81-104 to become associated with B cells in a stimulatory fashion, and this time is similar for live and paraformaldehyde-fixed B cells, which cannot internalize or process the peptide. Washed free of excess peptide after 2 h, B cells lose their ability to stimulate T cells by 8 to 12 h, with a time course indistinguishable from that for the loss of processed native Pc. Prolonged incubation of B cells with the peptide for 18 to 24 h results in a dramatic loss of the ability to present Pc 81-104. Even when provided with fresh Pc or Pc 81-104, these cells have diminished ability to present these Ag. This loss is selective, inasmuch as these B cells remain equivalent to untreated B cells in the presentation of an unrelated Ag, OVA, to an I-Ak-restricted specific T cell. However, the ability to present another I-Ek-restricted antigenic peptide of the D glycoprotein of HSV to its specific T cell is also diminished. Loss of activity is observed after incubation only with the peptide and not with the native protein and is not due to a depletion of the antigenic peptide from the incubation medium.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1988

Receptor-mediated B cell antigen processing. Increased antigenicity of a globular protein covalently coupled to antibodies specific for B cell surface structures.

Helper T cell recognition of globular protein antigens requires the intracellular processing of the native molecule by an antigen-presenting cell and subsequent presentation of a peptide fragment, containing the antigenic determinant, on the cell surface where it is recognized by the specific T cell in conjunction with Ia. B lymphocytes can function as antigen-presenting cells and, when antigen is bound by their surface Ig, are greatly enhanced in this capacity. In this report it is demonstrated that pigeon cytochrome c covalently coupled to antibodies directed toward either B cell surface immunoglobulin, class I or class II are effectively processed and presented by B cells to cytochrome c-specific T cells, requiring up to 1000-fold less cytochrome c as compared with cytochrome c alone or cytochrome c coupled to nonspecific immunoglobulin. The potent activity of the cytochrome c-antibody conjugates appears to be due to the ability of B cells to concentrate the antigen when the process becomes receptor mediated rather than to a signal provided to the B cell by the conjugate binding, because cytochrome c was not more effectively presented in the presence of unconjugated antibodies as compared with cytochrome c alone. Furthermore, the binding of the native antigen to B cell surfaces is not alone sufficient for T cell activation, in that the cytochrome c-antibody conjugates require processing and are major histocompatibility complex restricted. The results presented here indicate that surface immunoglobulin is not unique in its ability to facilitate antigen processing and/or presentation and that Ig, class I and class II are capable of transporting the cytochrome c to a cytoplasmic vesicle where proteolysis occurs yielding the required peptide, minimally of 10 amino acids. Cytochrome c coupled to monovalent fragments of anti-Ig-antibodies was nearly as effectively presented as cytochrome c coupled to bivalent antibodies, indicating that phenomena mediated by bivalent binding, such as patching and capping of the surface Ig, were not required for effective antigen presentation. The cytochrome c-antibody conjugates, which allow antigen processing to be initiated by receptor-mediated endocytosis, may provide the necessary tools to unravel the intracellular processes by which protein antigens are processed and presented by B lymphocytes.

Animals↗
PubMed1988

A Ca2+ influx in response to hypo-osmotic stress may alter osmolyte permeability by a phenothiazine-sensitive mechanism.

The phenomenon of cell volume recovery following a hypo-osmotic stress mediated by intracellular osmolyte regulation is well known. In many, perhaps all, cell types, the osmolytes involved are usually inorganic ions and amino acids. The details of the regulatory mechanisms for the organic-type osmolytes are not well known. We have found that an immediate influx of external Ca2+ occurs coincident with the application of a hypo-osmotic stress into red cells of two invertebrate species. In both, the influx is initiated by the osmotic stress, not the concomitant ionic decrease. Volume recovery in clam red blood cells is blocked by phenothiazines. In addition, the effect of the phenothiazines is to reduce the amino acid efflux; the ionic portion of the volume response is unaffected. In contrast, the phenothiazines potentiate the volume recovery in worm red coelomocytes. A23187 also potentiates the volume recovery of the worm red cells. The results suggest that the Ca2+ influx is involved in the mechanism that alters cell membrane permeability permitting the amino acid efflux by a mechanism that may involve calmodulin.

Amino Acids↗
PubMed1988

Identification and function of octopamine and tyramine conjugates in the Limulus visual system.

Major metabolites of octopamine and tyramine in the Limulus nervous system are identified here as gamma-glutamyl octopamine and gamma-glutamyl tyramine. We show that these conjugates are normal products of amine metabolism in Limulus, and that they are normally present in octopamine-rich Limulus tissues. The synthesis of these conjugates is not restricted to nervous tissue, but the highest activity of gamma-glutamyl amine synthetase was measured in the CNS. Our interest in these molecules stems from our previous observations which showed that they were synthesized and stored in, and released from, the efferent fibers to Limulus eyes which modulate the sensitivity of the eyes to light. Here we provide direct evidence for the release of the conjugates from Limulus eyes in response to depolarization, and that gamma-glutamyl octopamine can increase the sensitivity of the lateral eye to light. Our observations lend support to the hypothesis that gamma-glutamyl octopamine may serve as an intercellular messenger in the Limulus visual system.

Animals↗
PubMed1988

Antigen-presenting function of B lymphocytes.

Here we review our current results studying B cells as APC and the mechanisms by which processed antigen is transported to and held on the cell surface for recognition by the specific T cell along with the MHC class II molecules. These studies were carried out using the globular protein cytochrome c as antigen for which the T-cell antigenic determinant was localized to a C-terminal 10-amino acid peptide fragment. For certain analyses, native cytochrome c or antigenic peptide fragments were covalently coupled to antibodies directed toward B-cell surface structures, allowing the targeting of antigen to the APC surface. Our findings indicate that all B cells function as APC and that the APC function is not differentially regulated in defined B-cell subpopulations. Using cytochrome c-antibody conjugates, it was shown that the surface Ig plays two significant roles in augmenting the B-cell APC function following antigen binding: signalling for enhanced APC function and concentrating antigen for subsequent internalization and processing. Both IgM and IgD appear to function identically in facilitating antigen processing in both immune and nonimmune B-cell populations. Furthermore, the surface Ig does not appear to be specially differentiated to function in concentrating antigen, as antigen artificially bound to other B-cell surface structures including MHC class I and class II molecules is also effectively presented. Lastly, evidence is presented that a previously described B-cell activating factor activity is strongly associated with the membranes of activated but not unactivated helper T cells, providing a mechanism by which the T-cell helper function can be focused on the specific antigen-presenting B cell. Concerning the mechanism by which processed antigen is presented at the B-cell surface, evidence is presented suggesting a role of peptide-binding chaperone proteins which may function to transport peptide to the APC surface and facilitate its association with the appropriate Ia. One candidate protein, PBP72/74, is described which binds peptides but not native antigens, is a member of the hsp70 family and appears to play a role in antigen presentation by the ability of antisera raised against it to block APC functions. Peptide-antibody conjugates were used to explore the spacial restrictions on MHC-restricted peptide presentation and it was shown that peptides covalently coupled to antibodies specific for Ig, class I or class II molecules are effective antigens in vitro even in the absence of processing.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

T cell activation by processed antigen is equally blocked by I-E and I-A-restricted immunodominant peptides.

The T cell response to a soluble protein requires the processing of the native antigen by an antigen-presenting cell (APC) to a peptide containing an antigenic determinant, which is transported to and bound on the antigen-presenting cell surface, where it is subsequently recognized by the specific T cell in the context of the appropriate Ia molecule. Investigating the response of a pigeon cytochrome c-specific, I-Ek-restricted T cell hybrid, which recognizes a determinant present within a 10-amino acid C-terminal fragment of the protein, it was previously demonstrated that peptides homologous to the peptide from pigeon cytochrome c, but which were not stimulatory, blocked the T cell response to pigeon cytochrome c as processed and presented by APC. In this report the ability of a series of fourteen, 20-amino acid overlapping peptides, representing the entire length of staphylococcal nuclease (Nase), were assessed for their ability to block the response of a pigeon cytochrome c-specific T cell hybrid to antigen-pulsed presenting cells. Only three Nase peptides blocked the I-Ek-restricted pigeon cytochrome c-specific T cell response. Two of these, Nase 61-80 and Nase 91-110, function as T cell antigens in the I-Ad and I-Ab-restricted response to Nase. The third blocking peptide, Nase 101-120, has not been shown to be a T cell antigen. Two other peptides, Nase 51-70 and Nase 81-100, which are recognized by Nase-specific T cells in the context of I-Ek, have no effect on the I-Ek-restricted cytochrome c-specific T cell response. None of these peptides block the higher affinity, heteroclitic response of pigeon cytochrome c-specific T cells to tobacco hornworm moth cytochrome c. Moreover, the response of an I-Ak-restricted T cell to ovalbumin was blocked by the I-Ek-restricted cytochrome c peptides from three different species. Thus, peptides with no obvious primary amino acid sequence homology, and which are not capable of being recognized in the context of the same Ia, compete with one another for the sites on the APC necessary for presentation of processed antigen to T cells. These results suggest that there are structures on the APC surface in addition to Ia, which are necessary for effective antigen presentation following processing. One suitable candidate for such a cell surface material is the recently identified peptide-binding protein, PBP72/74 (Lakey et al., Proc. Natl. Acad. Sci. USA 1987. 84: 1659).

Animals↗
PubMed1987

Identification of a peptide binding protein that plays a role in antigen presentation.

The helper T-cell response to globular proteins appears, in general, to require intracellular processing of the antigen, such that a peptide fragment containing the T-cell antigenic determinant is released and transported to and held on the surface of an Ia-expressing, antigen-presenting cell. However, the molecular details underlying these phenomena are largely unknown. The means by which antigenic peptides are anchored on the antigen-presenting cell surface was investigated. A cell surface protein is identified that was isolated by its ability to bind to a 24-amino acid peptide fragment of pigeon cytochrome c, residues 81-104, containing the major antigenic determinant for B10.A mouse T cells. This peptide binding protein, purified from [35S]methionine-labeled cells, appears as two discrete bands of approximately equal to 72 and 74 kDa after NaDodSO4/PAGE. The protein can be eluted from the peptide affinity column with equivalent concentrations of either the antigenic pigeon cytochrome c peptide or the corresponding nonantigenic peptide of mouse cytochrome c. However, it does not bind to the native cytochromes c, either of pigeon or mouse, and thus the protein appears to recognize some structure available only in the free peptides. This protein plays a role in antigen presentation as evidenced by the ability of rabbit antibodies raised against it to block the activation of an antigen-specific T-cell hybrid by antigen-presenting cells and pigeon cytochrome c. Its expression is not major histocompatibility complex-restricted in that the blocking activity of the antisera can be absorbed on spleen cells from mice of different haplotypes. This peptide binding protein can be isolated from a variety of cell types, including B cells, T cells, and fibroblasts. The anchoring of processed peptides on the cell surface by such a protein may play a role in antigen presentation--facilitating the interaction of antigenic peptides with Ia and/or the T-cell receptor.

Animals↗
PubMed1986

Role of L3T4 and Ia in the heteroclitic response of T cells to cytochrome c.

The activation of helper T lymphocytes has been proposed to result from the sum of low-affinity interactions between the specific immune receptor, as well as nonpolymorphic receptors such as L3T4 on the T cell surface, and nominal antigen and Ia displayed in a multivalent array on the antigen-presenting cell surface. The present work takes advantage of a T cell hybridoma specific for pigeon cytochrome c in the context of I-Ek, which responds to tobacco hornworm moth cytochrome c at one hundredth the concentration of the homologous antigen, to determine if the T cell's requirement for L3T4 and Ia is directly related to its functional affinity for antigen. The results demonstrate that the T cell's activation by pigeon cytochrome c was blocked by antibodies directed to L3T4 and to I-Ek, even at antigen concentrations twofold to fourfold above those required for maximal responses. In contrast, the response to tobacco hornworm moth cytochrome c was not as affected by these antibodies under equivalent superoptimal conditions. The same phenomenon was observed for the T cell's activation by the carboxyl-terminal peptide fragments of the two cytochromes c, which do not require processing, indicating that the differences were not due to the relative efficiency of processing and/or presentation of the antigens. Although both I-Ek- and L3T4-specific antibodies blocked the T cell response to pigeon cytochrome, antibodies to I-Ak had no effect, even though I-Ak had been considered to be a ligand for L3T4. Thus, either Ia does not bind L3T4 or, if it does, I-Ek must be a sufficient ligand for L3T4 for T cells that recognize their antigen in the context of I-Ek. These studies provide more definitive evidence that the T cell's requirement for the functions of Ia and of L3T4 is dependent on the T cell's functional affinity for its antigenic determinant. This data is consistent with a model of T cell activation in which, given a high enough affinity of the T cell receptor for the processed antigen, the requirement for other components of a stimulatory complex, such as Ia and L3T4, may diminish to undetectable levels.

Animals↗
PubMed1986

Antigen presentation is a function of all B cell subpopulations separated on the basis of size.

Purified splenic B cells from nonimmune mice were separated by counterflow centrifugal elutriation into 6 subpopulations containing cells of discrete sizes ranging from 119 to 200 micron3. B cells of each subpopulation were competent to process and present a native globular protein antigen, cytochrome c, to a cytochrome c-specific T cell hybrid. In all cases, the B cells' antigen-presenting function was radiation sensitive and did not require T cells or T cell products, since B cells fixed with paraformaldehyde effectively presented a carboxyl-terminal peptide fragment of cytochrome c containing the T cell determinant. Furthermore, the antigen-presenting function of B cells of each subpopulation was augmented by treatment with submitogenic doses of the F(ab')2 fragment of rabbit anti-mouse Ig antibodies, in that 10-30-fold fewer B cells were required and higher maximal T cell responses were achieved, indicating that B cells of all sizes are capable of being regulated in their antigen presentation function through their surface Ig. In addition, B cells of each subpopulation responded to soluble factors present in the supernatants of activated T cells as evidenced by an increase in volume and by the uptake of [3H]thymidine. These results indicate that B cells, regardless of size, are able to participate in at least two essential phases of T cell-dependent antibody responses, initiating the interaction by processing and presenting antigen to helper T cells and responding to soluble helper factors secreted by activated T cells.

Animals↗
PubMed1986

Peptides related to the antigenic determinant block T cell recognition of the native protein as processed by antigen-presenting cells.

A mouse T cell hybrid specific for pigeon cytochrome c in the context of I-Ek responds by secreting interleukin 2 when co-cultured with the native antigen and the B cell lymphoma, LK-35.2, or naive splenic B cells as antigen-presenting cells (APC). Cytochromes c and their corresponding C-terminal fragments which are not capable of stimulating the TPc9.1 cells, including the autologous mouse cytochrome c, block the T cells' response to pigeon cytochrome c. In contrast, nonstimulatory N-terminal peptides of cytochrome c, which share no homology with the antigenic peptide, do not block. Blocking is observed when the nonstimulatory cytochromes c or peptides are present in culture with the live APC and nonsaturating concentrations of pigeon cytochrome c. With tobacco hornworm moth cytochrome c as antigen, a protein for which the T cell has a higher functional affinity, the response of TPc9.1 cannot be blocked by the nonstimulatory cytochromes c or by peptides, even when limiting concentrations of the tobacco hornworm moth cytochrome c are used. When paraformaldehyde-fixed APC are employed, no native cytochrome c can stimulate the T cells, including the tobacco hornworm moth protein which with the live APC is effective at 50 to 100-fold lower concentrations than pigeon cytochrome c. However, with fixed APC the T cells are stimulated by the C-terminal fragments containing residues 81-104 of the pigeon protein or residues 81-103 of the tobacco hornworm moth protein as readily and with the same relative efficiencies as the native protein, presented by live APC. The nonstimulatory peptides, but not the native cytochromes c, block T cell activation by pigeon cytochrome c pulsed-fixed APC, indicating that the nonstimulatory peptides compete with the stimulatory pigeon cytochrome c peptides produced by the APC. This competition appears to be due to nonstimulatory peptides which associate at the APC surface and not to those acting from solution because the APC which have been incubated with pigeon cytochrome c and nonstimulatory peptides and washed free of excess antigen and peptides are not stimulatory to the T cell hybrid. It was concluded that the activation of a pigeon cytochrome c-specific T cell, which recognizes a peptide fragment of the native protein on the surface of an APC, can be blocked by an excess of nonstimulatory homologous peptides when these are also associated on the surface of the APC.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1986

Octopamine potentiates intracellular Na+ and Cl- reductions during cell volume regulation in Limulus exposed to hypoosmotic stress.

The biogenic amine octopamine (OCT) appears to be involved in cell volume regulation in the horseshoe crab, Limulus polyphemus, during hypoosmotic stress. OCT is present in relatively large amounts (160 nmoles/g dry wt) in the cardiac ganglion. Furthermore, OCT is released from the isolated ganglion during exposure to hypoosmotic media. This release is reflected in the elevation of blood OCT concentrations from basal levels of 4 X 10(-9) M reaching 1.2 X 10(-8) M within 72 h of exposure of animals to hypoosmotic media. The circulating OCT potentiates the hypoosmotically-induced reductions of intracellular Na+ and Cl- by a ouabain-sensitive mechanism which complements the main ion regulating (ouabain-insensitive) mechanisms utilized during cell volume recovery.

Animals↗
PubMed1985

Comparative electrical properties of identified neurons in Elysia chlorotica before and after low salinity acclimation.

Identified neurons in the abdominal ganglion of Elysia chlorotica adapted to 50% seawater (SW) had significantly different electrical properties from the same cells in animals adapted to 100% SW. Resting potential, action potential (AP) overshoot, (AP) duration, threshold and after potential were all different following salinity acclimation. The resting potential of these cells behaves as an ideal potassium electrode above 10 mM [K+]. The action potential has both sodium and calcium components to the rising phase.

Acclimatization↗
PubMed1985

Anti-immunoglobulin augments the B-cell antigen-presentation function independently of internalization of receptor-antigen complex.

All mouse splenic B cells, including small resting B cells, process and present the native globular protein antigens, pigeon and tobacco hornworm moth cytochromes c, to a cytochrome c-specific T-cell hybrid in a major histocompatibility complex-restricted fashion, in the micromolar to nanomolar antigen-concentration range. As is the case for macrophages, treatment with paraformaldehyde or the lysosomotropic agents chloroquine and ammonium chloride blocked processing of the native pigeon protein but did not affect the presentation of a carboxyl-terminal peptide fragment of pigeon cytochrome c (residues 81-104) which contained the T-cell antigenic determinant. However, in contrast to macrophages, whose antigen-processing and -presentation functions are insensitive to radiation, radiation blocked the processing of the native protein but not the presentation of the peptide fragment. The processing and presentation function of the B cells was augmented by F(ab')2 of rabbit anti-mouse Ig antibodies, in that 1/10th to 1/30th as many cells and 1/10th as much antigen were required to maximally activate the T-cell hybrid. This augmentation did not appear to be due to either crosslinking of the Ig receptors or to B-cell activation, as monovalent Fab fragments were nearly as effective as the bivalent reagent, and the concentrations of F(ab')2 anti-Ig used did not induce measurable proliferative responses. Furthermore, enhancement can occur in the absence of cytochrome c binding and internalization, since B cells that were fixed with paraformaldehyde after treatment with F(ab')2 anti-Ig were more effective in presenting the carboxyl-terminal peptide than were untreated fixed cells. The same phenomenon followed the binding of an irrelevant antigen (carboxydinitrophenylated bovine serum albumin) by antigen-binding B cells, resulting in enhanced processing and/or presentation of native pigeon cytochrome c. Thus, nonspecific enhancement of antigen processing and presentation can be obtained by either antigen or anti-Ig binding to the B-cell antigen receptor, both treatments presumably delivering the same signal without requiring internalization of the specifically bound antigen for subsequent processing.

Animals↗