Optimization of antigen presentation to T cell hybridomas by purified Ia molecules in planar membranes

Quill, Helen; Carlson, Letitia; Fox, Barbara S.; Weinstein, John N.; Schwartz, Ronald H.

doi:10.1016/0022-1759(87)90432-7

Cited by 17 publications

(5 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Lipids, by interacting directly with the MHC protein or by changing the structure ofthe detergent micelles, may induce conformational changes reflected in the distal binding site of the protein (23,24). Consistent with this hypothesis are the observations indicating that class II MHC molecules reconstituted in lipid monolayers present peptides differently, depending on the lipid composition (25). Other studies have shown that treatment of antigen-presenting cells with purified phospholipase A2 or C, or with cerulenin, an antibiotic that interferes with lipid metabolism, greatly diminished their antigen-presenting function (26)(27)(28).…”

mentioning

confidence: 69%

Phospholipids enhance the binding of peptides to class II major histocompatibility molecules.

Roof

Luescher²,

Unanue³

1990

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

The binding of a lysozyme and ovalbumin peptide to purified class II major histocompatibiity molecules in detergents was increased by the addition of certain lipids. Natural lipids from B lymphoma cells enhanced the binding and so did phosphatidylcholine, phosphatidylserine, phosphatidylinositol, and cardiolipin. Phosphatidylethanolamine, sphingomyelin, and cholesterol had no effect. There was no major difference between the effects of a phospholipid and its lyso derivative. As studied with phosphatidylcholine, the increase in peptide binding was also dependent on the fatty acid composition of the lipid. (MHC) proteins (1,2). The extent of binding of peptides to histocompatibility molecules, which can be studied in detergent solutions (1-7), usually correlates with their immunogenicity in vivo (2-4). There are, however, indications that the interactions of peptides with class II MHC may be quantitatively different on live antigen-presenting cells. For example, the binding of antigenic peptides to class II MHC molecules on live cells was reported to be much faster than that found in solution (8-10). A similar result was found for the dissociation of peptides bound to the class II MHC molecules (11).We recently found that the labeling of class II MHC molecules to peptides containing a photoreactive compound was influenced by phospholipids, both in the extent as well as in the relative degree ofthe a or p chain labeling (12). When tested on cell membranes that contained class II MHC proteins, photoreactive conjugates of the hen egg white lysozyme (HEL) peptide from residues 46 to 61 labeled exclusively the a chain of I-Ak (13). Conversely, I-Ak purified in an equimolar solution of MEGA 8 and MEGA 9 detergents was labeled to similar extent on both chains. However, the addition of certain lysophospholipids resulted in strong binding, with lysophosphatidylcholine predominantly labeling the a chain and lysophosphatidylserine labeling the ,B chain. We concluded that class II MHC molecules contained dialyzable components required for their binding integrity and that, most likely, these components were lipids.Lipid-protein interactions are of critical importance in maintaining the biological activity of a number of membrane proteins (for review, see refs. 14-16). These interactions may have different degrees of specificity and often induce conformational changes in the protein critical for their function (14,(17)(18)(19)(20)(21)(22). Lipids, by interacting directly with the MHC protein or by changing the structure ofthe detergent micelles, may induce conformational changes reflected in the distal binding site of the protein (23,24). Consistent with this hypothesis are the observations indicating that class II MHC molecules reconstituted in lipid monolayers present peptides differently, depending on the lipid composition (25). Other studies have shown that treatment of antigen-presenting cells with purified phospholipase A2 or C, or with cerulenin, an antibiotic that interferes with lipid metabolism, greatly dimi...

show abstract

mentioning

confidence: 69%

Phospholipids enhance the binding of peptides to class II major histocompatibility molecules.

Roof

Luescher²,

Unanue³

1990

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…Purified class II was reconstituted into liposomes from solutions containing 1% deoxycholate, 275/~M phospholipid, 75/~M cholesterol, and 1/~M class II by extensive dialysis against PBS. The phospholipids used were t-ot-dipalmitoyl phosphatidylcholine and t-c~-dilindeoyl phosphatidylcholine (Sigma Chemical Co., St. Louis, MO) in a 9:1 molar ratio (11). Liposomes were supported by incubation with 2.5 x 106 washed 10-#m silica gel particles (Sigma Chemical Co.) for 2 h at 24~ Liposome-coated particles were washed and resuspended in buffer immediately before use.…”

Section: Methodsmentioning

confidence: 99%

“…OVA(323-339) (18,19) and Myo(106-118) (20,21) are known to bind to I-A a, whereas HEL(104-120) binds bE d (7,22). HEL(46-61) (23,24) and pCC(91-104) (11,25) have very low a~nity for these class II molecules. Myo(106-118) partially inhibited the binding of biotin/peptide to I-E a at higher concentrations, although it is not known as a T cell determinant recognized in association with I-A d. A similar result has been reported by Sette et al (26).…”

Section: Accessory Molecules Are Not Required For Enhanced Peptide/clmentioning

confidence: 99%

Enhanced binding of peptide antigen to purified class II major histocompatibility glycoproteins at acidic pH.

Jensen

1991

The Journal of Experimental Medicine

View full text Add to dashboard Cite

SllmmsryHelper T lymphocytes recognize peptide antigens stably associated with class II major histocompatibility complex (MHC) glycoproteins on the surface of antigen-presenting cells and serve to regulate a wide variety of immune responses. A previous study from our laboratory had demonstrated that the functional association of various peptide antigens with the antigen-presenting cell membrane was increased at pH 5 as compared to pH 7, consistent with the potential role of acidic endosomal compartments in antigen processing. The mechanism for this effect was not determined. In the present study, assays using purified class II glycoprotein were used to further define this mechanism. The potential requirement for pH-dependent interactions involving non-MHC membrane components was excluded in functional assays with purified class II reconstituted in artificial membranes containing only neutral phospholipids and cholesterol. The association of HEL(104-120) with I-E a, and OVA(323-339) with I-A a, was increased at pH 5, as measured by activation of specific T cell hybridomas. An enzyme immunoassay was developed to measure the binding of biotin-labeled peptides to purified class II in detergent micelles. The pH dependence of binding paralleled our previous functional results. Optimum binding of biotin-HEL(104-120) to I-E d was observed at pH "~4.5, whereas maximum binding of biotin-Myo(106-118) to I-A d occurred at pH ~5.5. The latter peptide also bound weakly to I-E d, but with a pH dependence similar to that observed using HEL(104-120). Further experiments with biotin-HEL(104-120)/I-E d indicated that both the apparent af~nity and the apparent concentration of peptide-binding sites are increased as hydrogen ion concentration is increased from pH 7 to pH 5. The effect of pH in this range was largely reversible and was not associated with a change in peptide dissociation that could be measured with our assay system. Binding was not inhibited in the presence of 1.5 M NaC1, suggesting that electrostatic interactions between HEL(104-120) and I-E d are not essential for binding. It is proposed that protonation of a critical group(s) in the class II molecule regulates its capacity to form stable complexes with peptide. However, this effect alone does not fully account for the rapid kinetics ofpeptide binding observed in experiments with intact antigen-presenting cells.

show abstract

“…2B4.11, murine T cell hybridoma cells [10] and LK 35.2 (B cell hybridoma) cells were cultured in RPMI 1640 medium (Gibco, Mulgrave, Australia) supplemented with 10% foetal calf serum (FCS), 2mM glutamine, 50U/ml penicillin/streptomycin and 0.002% mercaptoethanol at 37ºC and 5% CO 2 . LK 35.2 cells were used to present cytochrome c fragments to 2B4.11 cells [11].…”

Section: Cells and Dna Constructsmentioning

confidence: 99%

T-Cell Antigen Receptor Assembly and Cell Surface Expression Is Not Affected by Treatment with T-Cell Antigen Receptor-Alpha Chain Transmembrane Peptide

Kurosaka

Bolte²,

Ali³

et al. 2007

PPL

View full text Add to dashboard Cite

A synthetic peptide termed core peptide (CP), which corresponds to a specific sequence of the TCR-alpha chain transmembrane domain, is known to inhibit IL-2 production in antigen stimulated T-cells. The molecular mechanism of the TCR inhibition is not known. This study examined the effects of CP on TCR subunit assembly and TCR cell surface expression in vitro. Co-transfection experiments between TCR-alpha and CD3-delta using COS-7 cells, and the interaction between TCR-alpha and the CD3 proteins in a T-cell line (2B4) were analysed after incubation with CP or its conjugates. Results indicate that CP co-precipitates with CD3-delta and CD3-epsilon in vitro, without any effect on TCR-alpha/CD3-delta dimerisation or TCR multisubunit assembly and cell surface expression.

show abstract

Optimization of antigen presentation to T cell hybridomas by purified Ia molecules in planar membranes

Cited by 17 publications

References 23 publications

Phospholipids enhance the binding of peptides to class II major histocompatibility molecules.

Phospholipids enhance the binding of peptides to class II major histocompatibility molecules.

Enhanced binding of peptide antigen to purified class II major histocompatibility glycoproteins at acidic pH.

T-Cell Antigen Receptor Assembly and Cell Surface Expression Is Not Affected by Treatment with T-Cell Antigen Receptor-Alpha Chain Transmembrane Peptide

Contact Info

Product

Resources

About