The MHC class II molecule H2-M is targeted to an endosomal compartment by a tyrosine-based targeting motif

Lindstedt, Ragnar; Liljedahl, Monika; Péleraux, Annick; Peterson, Per A.; Karlsson, Lars

doi:10.1016/1074-7613(95)90127-2

Cited by 87 publications

(64 citation statements)

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“…Mutagenesis of the HLA-DRB1*0101 cDNA was performed using a cassette-cloning strategy, overlap-extension PCR, or a combination of the two, as previously described (18,19). In brief, two cloning sites were introduced in the cDNA by PCR, flanking the region corresponding to interface region I in the ␤1 domain to facilitate the cassette-cloning strategy.…”

Section: Mutagenesis Of Hla-drb1*0101 Cdna and Construction Of Taggedmentioning

confidence: 99%

Amino Acid Substitutions in the Putative MHC Class II “Dimer of Dimers” Interface Inhibit CD4+ T Cell Activation

Lindstedt

Monk

Lombardi

et al. 2001

The Journal of Immunology

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Activation of T lymphocytes is dependent on multiple ligand-receptor interactions. The possibility that TCR dimerization contributes to T cell triggering was raised by the crystallographic analysis of MHC class II molecules. The MHC class II molecules associated as double dimers, and in such a way that two TCR (and two CD4 molecules) could bind simultaneously. Several subsequent studies have lent support to this concept, although the role of TCR cross-linking in T cell activation remains unclear. Using DRA cDNAs modified to encode two different C-terminal tags, no evidence of constitutive double dimer formation was obtained following immunoprecipitation and Western blotting from cells transiently transfected with wild-type DRB and tagged DRA constructs, together with invariant chain and HLA-DM. To determine whether MHC class II molecules contribute actively to TCR-dependent dimerization and consequent T cell activation, panels of HLA-DR1β and H2-Ek cDNAs were generated with mutations in the sequences encoding the interface regions of the MHC class II double dimer. Stable DAP.3 transfectants expressing these cDNAs were generated and characterized biochemically and functionally. Substitutions in either interface region I or III did not affect T cell activation, whereas combinations of amino acid substitutions in both regions led to substantial inhibition of proliferation or IL-2 secretion by human and murine T cells. Because the amino acid-substituted molecules were serologically indistinguishable from wild type, bound antigenic peptide with equal efficiency, and induced Ag-dependent CD25 expression indicating TCR recognition, the reduced ability of the mutants to induce full T cell activation is most likely the result of impaired double dimer formation. These data suggest that MHC class II molecules, due to their structural properties, actively contribute to TCR cross-linking.

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Section: Mutagenesis Of Hla-drb1*0101 Cdna and Construction Of Taggedmentioning

confidence: 99%

Amino Acid Substitutions in the Putative MHC Class II “Dimer of Dimers” Interface Inhibit CD4+ T Cell Activation

Lindstedt

Monk

Lombardi

et al. 2001

The Journal of Immunology

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“…These data, together with other published work (10,14), support the concept that the germ line-encoded regions of the TCR are inherently MHC-specific, emphasizing that complementarity to the peptide͞MHC-binding site is a major evolutionary selective pressure on the TCR. What is not clear from previous work is whether or not specific features of MHC molecules are required for interactions with TCRs.HLA-DM (DM) and the mouse homologue H-2M are predominantly sequestered in endosomal-like compartments known as MIICs (15) due to a tyrosine-based motif in the tail of the ␤ chain (16,17). This MHC-encoded molecule has strong tertiary structural similarity to MHC class II (18, 19), but, due to its intracellular residence, it has been removed from the forces of evolution that have been hypothesized to drive and maintain .…”

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confidence: 99%

“…HLA-DM (DM) and the mouse homologue H-2M are predominantly sequestered in endosomal-like compartments known as MIICs (15) due to a tyrosine-based motif in the tail of the ␤ chain (16,17). This MHC-encoded molecule has strong tertiary structural similarity to MHC class II (18, 19), but, due to its intracellular residence, it has been removed from the forces of evolution that have been hypothesized to drive and maintain .…”

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confidence: 99%

Coevolution of TCR-MHC interactions: Conserved MHC tertiary structure is not sufficient for interactions with the TCR

Kim

Guo

Sant’Angelo

2005

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

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The specificity for self-MHC that is necessary for T cell function is a consequence of intrathymic selection during which T cell antigen receptors (TCRs) expressed by immature thymocytes are tested for their affinity for self-peptide:self-MHC. The germ-line-encoded segments of the TCR, however, are believed to have an innate specificity for structural features of MHC molecules. We directly tested this hypothesis by generating a transgenic mouse system in which the protein HLA-DM is expressed at the surface of thymic cortical epithelial cells in the absence of classical MHC molecules. The specialized intracellular function of HLA-DM has removed this MHC class II-like protein from the evolutionary forces that have been hypothesized to shape TCR-MHC interactions. Our study shows that a structural mimic of MHC class II is not sufficient to appropriately interact with the TCRs expressed by developing thymocytes. This result emphasizes the unique complementarity of TCR-MHC interactions that are maintained by the evolutionary pressures dictated by positive selection.T cell receptor ͉ thymocytes ͉ mice ͉ T lymphocytes ͉ thymus D evelopment of thymocytes is a multistep process that shapes the repertoire of mature T cells. Ideally, the mature T cell antigen receptor (TCR) repertoire is depleted of specificities that are overtly self-reactive, but is also diversified enough to mount effective immune responses (1, 2). Selection is based on the interactions of the TCRs expressed by immature CD4 ϩ CD8 ϩ , double-positive (DP) thymocytes with selfpeptide:self-MHC expressed on the surface of thymic stromal cells. Poorly defined criteria determine whether this receptorligand interaction is of sufficient avidity to drive development of DP thymocytes to mature single-positive, MHC class I-or MHC class II-restricted T cells.Unresolved issues regarding the positive selection of T cells include the extent that specific interactions of the TCR with MHC-bound self-peptides rather than interactions with MHC itself shape the mature CD4 ϩ T cell repertoire. This question has been addressed by a number of systems that have not produced concordant results (3-7). Such contradictory data have led to speculation that different TCRs may have different requirements for recognition of MHC-bound self-peptide. The inherent MHC reactivity of the preselection TCR repertoire is a second issue that has not been definitively answered. In other words, it is not clear whether the germ line-encoded regions of the TCR are predisposed toward interactions with MHC, or, alternatively, whether the ability of TCRs expressed by mature T cells to interact with MHC is fully a consequence of positive selection.Mutagenesis studies have suggested that the TCR has a conserved orientation of interaction with MHC molecules, and, therefore, conserved interactions between MHC molecules and TCRs seemed likely. Crystallographic studies of TCRs bound to MHC, however, have not fully confirmed this prediction (7). In retrospect, perhaps this finding is not surprising, consideri...

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“…3 shows that wild-type HLA-DM (B) accumulates in intracellular vesicles that are also positive for the lysosomal marker Lamp-1 (G). A tyrosine-based motif on the cytoplasmic tail of DM␤ is responsible for its accumulation in endocytic compartments (31)(32)(33). The three mutated forms of DM were also found in peripheral vesicles (Fig.…”

Section: Resultsmentioning

confidence: 99%

Functional Analysis of Tryptophans α62 and β120 on HLA-DM

Faubert¹,

Samaan²,

Thibodeau³

2002

Journal of Biological Chemistry

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In the endocytic pathway of antigen-presenting cells, HLA-DM catalyzes the exchange between class II-associated invariant chain peptide (CLIP) and antigenic peptides onto major histocompatibility complex class II molecules. At low pH of lysosomal compartments, both HLA-DM and HLA-DR undergo conformational changes, and it was recently postulated that two partially exposed tryptophans on HLA-DM might be involved in the interaction between the two molecules. To define contact regions on HLA-DM, we have conducted site-directed mutagenesis on those two hydrophobic residues. The HLA-DM ␣W62A,␤W120A (DM W62A/W120A ) double mutant was expressed in HLA-DR ؉ HeLa cells expressing invariant chain, and the activity of this DM molecule was assessed. Flow cytometry analysis of cell surface DR-CLIP complexes revealed that DM W62A/W120A removes CLIP as efficiently as its wild-type counterpart. DM W62A/W120A was found in the endocytic pathway by immunofluorescence, and DM-DR complexes were immunoprecipitated from these cells at pH 5. Finally, mutations ␣W62A and ␤W120A on HLA-DM did not affect the association with HLA-DO. The complex egresses the endoplasmic reticulum and accumulates in endocytic vesicles. Moreover, DO and DM W62A/W120A were co-immunoprecipitated at pH 7. We conclude that the ␣62 and ␤120 tryptophan residues are not required for the activity of DM, nor are they directly implicated in the interaction with DR or DO.Major histocompatibility complex class II molecules are primordial for activation of CD4ϩ T cells and for immunological response against pathogens (1). Three major histocompatibility complex II ␣/␤ heterodimers associate with a trimer of invariant chain (Ii) 1 to form a nanomeric complex in the endoplasmic reticulum (ER) (2). This association allows the proper folding and trafficking of the major histocompatibility complex II molecules (3-5). Gradual proteolysis of Ii occurs in the endocytic compartments by the sequential action of proteases including cathepsins (6). A residual peptide of Ii (CLIP) remains in the peptide groove of the class II molecule and stabilizes the heterodimer by preventing collapse of the groove (7,8). For most allotypes, CLIP must be actively removed to allow binding of antigenic peptides. HLA-DM, a nonclassical intracellular class II molecule, plays a central role in the efficiency of antigen presentation as it catalyzes CLIP release from HLA-DR and stabilizes the class II in a chaperone-like fashion (9 -14). In fact, mice lacking the H2-Ma gene express a high level of surface class II-CLIP complexes (15-17).The precise molecular mechanism of action of HLA-DM remains to be established. X-ray diffraction studies on HLA-DM crystals revealed a closed peptide-binding groove and an overall quaternary structure similar to classical class II molecules (18,19). Time-resolved fluorescence anisotropy and far-UV circular dichroism spectrum analysis using soluble HLA-DM revealed that its structure is quite rigid and that it is not subjected to gross pH-dependent conformational change. Ho...

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The MHC class II molecule H2-M is targeted to an endosomal compartment by a tyrosine-based targeting motif

Cited by 87 publications

References 50 publications

Amino Acid Substitutions in the Putative MHC Class II “Dimer of Dimers” Interface Inhibit CD4+ T Cell Activation

Amino Acid Substitutions in the Putative MHC Class II “Dimer of Dimers” Interface Inhibit CD4+ T Cell Activation

Coevolution of TCR-MHC interactions: Conserved MHC tertiary structure is not sufficient for interactions with the TCR

Functional Analysis of Tryptophans α62 and β120 on HLA-DM

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