Inhibitory Fragment from the p41 Form of Invariant Chain Can Regulate Activity of Cysteine Cathepsins in Antigen Presentation (original) (raw)
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The p41 isoform of invariant chain is a chaperone for cathepsin L
The EMBO journal, 2001
The p41 splice variant of major histocompatibility complex (MHC) class II-associated invariant chain (Ii) contains a 65 aa segment that binds to the active site of cathepsin L (CatL), a lysosomal cysteine protease involved in MHC class II-restricted antigen presentation. This segment is absent from the predominant form of Ii, p31. Here we document the in vivo significance of the p41-CatL interaction. By biochemical means and electron microscopy, we demonstrate that the levels of active CatL are strongly reduced in bone marrow-derived antigen-presenting cells that lack p41. This defect mainly concerns the mature two-chain forms of CatL, which depend on p41 to be expressed at wild-type levels. Indeed, pulse-chase analysis suggests that these mature forms of CatL are degraded by endocytic proteases when p41 is absent. We conclude that p41 is required for activity of CatL by stabilizing the mature forms of the enzyme. This suggests that p41 is not merely an inhibitor of CatL enzymatic a...
1996
is the proteolytic destruction of Ii, as intact ␣Ii trimers and Harold A. Chapman* themselves are unable to bind peptides (Roche and *Department of Medicine Cresswell, 1990). In vivo, inhibition of all cysteine class Brigham and Women's Hospital proteases impairs Ii breakdown and induces accumulaand Harvard Medical School tion of class II-associated Ii fragments in B lymphoblas-Boston, Massachusetts 02115 toid cells (Blum and Cresswell, 1988; Nguyen et al., 1988; † Center for Cancer Research Newcomb and Cresswell, 1993). Consequently, acquisi-Department of Biology tion of antigenic peptide by class II ␣ dimers is pre-Massachusetts Institute of Technology vented (Neefjes and Ploegh, 1992), expression of class Cambridge, Massachusetts 02139 II molecules at the cell surface is decreased (Neefjes ‡ Arris Pharmaceuticals, Incorporated and Ploegh, 1992; Bé naroch et al., 1995), and antigen-385 Oyster Point Boulevard stimulated T cell proliferation is attenuated (Buus and South San Francisco, California 94080 Werdelin, 1986; Diment, 1990). Despite the absolute requirement for Ii destruction to render class II molecules capable of binding peptide, the primary proteases that perform this task remain unidentified. Previous attempts Summary at identifying these proteases are difficult to interpret, owing to the rather nonspecific action of the inhibitors Destruction of Ii by proteolysis is required for MHC used and possible contamination of commercially availclass II molecules to bind antigenic peptides, and for able protease preparations with other proteases. For transport of the resulting complexes to the cell surexample, leupeptin, a commonly utilized protease inhibiface. The cysteine protease cathepsin S is highly tor (Nguyen et al., 1988), acts on many proteases of the expressed in spleen, lymphocytes, monocytes, and cysteine and serine class. Lysosomotropic agents, such other class II-positive cells, and is inducible with as quinidine (Humbert et al., 1993) and concanamycin interferon-␥. Specific inhibition of cathepsin S in B B (Bé naroch et al., 1995), neutralize acidic compartlymphoblastoid cells prevented complete proteolysis ments in a nonspecific manner. Also, analysis of some of Ii, resulting in accumulation of a class II-associated commercially available cathepsin B preparations in our 13 kDa Ii fragment in vivo. Consequently, the formation laboratory have demonstrated the presence of addiof SDS-stable complexes was markedly reduced. Puritional cysteine proteases (H. A. C., unpublished data). fied cathepsin S, but not cathepsin B, H, or D, specifi-Cathepsin S, a cysteine protease originally cloned cally digested Ii from ␣Ii trimers, generating ␣-CLIP from human alveolar macrophages, is highly expressed complexes capable of binding exogenously added in the spleen and professional antigen-presenting cells, peptide in vitro. Thus, cathepsin S is essential in B including B lymphocytes, macrophages, and other class cells for effective Ii proteolysis necessary to render II-positive cells (Shi et al., 1992, 1994; Morton et al., class II molecules competent for binding peptides. 1995). Moreover, it is inducible with interferon-␥ (IFN␥), it is a potent endoprotease, and it has a broad pH activity profile (Shi et al.
European Journal of Immunology, 1992
Proteolytic degradation (processing) of antigen by antigen-presenting cells is a major regulatory step in the activation of a T lymphocyte immune response. However, the enzymes responsible for antigen processing remain largely undefined. In this study we show that cathepsin E , and not the ubiquitous lysosomal cathepsin D, is the major aspartic proteinase in a murine antigenpresenting cell line, A20. This enzyme is localized to a non-lysosomal compartment of the endosomal system in these cells. Functional studies using a highly specific inhibitor of cathepsin E show that this enzyme is essential for the processing of ovalbumin by this cell line. Thus, cathepsin E, whose function was hitherto unknown, may play a major role in antigen processing.
Cathepsin S activity regulates antigen presentation and immunity
Journal of Clinical Investigation, 1998
MHC class II molecules display antigenic peptides on cell surfaces for recognition by CD4 ϩ T cells. Proteolysis is required in this process both for degradation of invariant chain (Ii) from class II-Ii complexes to allow subsequent binding of peptides, and for generation of the antigenic peptides. The cysteine endoprotease, cathepsin S, mediates Ii degradation in human and mouse antigen-presenting cells.
The EMBO Journal, 1999
The lysosomal cysteine proteases cathepsins S and L play crucial roles in the degradation of the invariant chain during maturation of MHC class II molecules and antigen processing. The p41 form of the invariant chain includes a fragment which specifically inhibits cathepsin L but not S. The crystal structure of the p41 fragment, a homologue of the thyroglobulin type-1 domains, has been determined at 2.0 Å resolution in complex with cathepsin L. The structure of the p41 fragment demonstrates a novel fold, consisting of two subdomains, each stabilized by disulfide bridges. The first subdomain is an α-helix-β-strand arrangement, whereas the second subdomain has a predominantly β-strand arrangement. The wedge shape and three-loop arrangement of the p41 fragment bound to the active site cleft of cathepsin L are reminiscent of the inhibitory edge of cystatins, thus demonstrating the first example of convergent evolution observed in cysteine protease inhibitors. However, the different fold of the p41 fragment results in additional contacts with the top of the R-domain of the enzymes, which defines the specificity-determining S2 and S1Ј substrate-binding sites. This enables inhibitors based on the thyroglobulin type-1 domain fold, in contrast to the rather non-selective cystatins, to exhibit specificity for their target enzymes.
The Journal of Experimental Medicine, 2000
The major histocompatibility complex (MHC) class II–associated invariant chain (Ii) regulates intracellular trafficking and peptide loading of MHC class II molecules. Such loading occurs after endosomal degradation of the invariant chain to a ∼3-kD peptide termed CLIP (class II–associated invariant chain peptide). Cathepsins L and S have both been implicated in degradation of Ii to CLIP in thymus and peripheral lymphoid organs, respectively. However, macrophages from mice deficient in both cathepsins S and L can process Ii and load peptides onto MHC class II dimers normally. Both processes are blocked by a cysteine protease inhibitor, indicating the involvement of an additional Ii-processing enzyme(s). Comparison of cysteine proteases expressed by macrophages with those found in splenocytes and dendritic cells revealed two enzymes expressed exclusively in macrophages, cathepsins Z and F. Recombinant cathepsin Z did not generate CLIP from Ii–MHC class II complexes, whereas cathepsin ...
Immunology, 2010
The expression of major histocompatibility complex class II (MHC II) molecules is post-translationally regulated by endocytic protein turnover. Here, we identified the serine protease cathepsin G (CatG) as an MHC II-degrading protease by in vitro screening and examined its role in MHC II turnover in vivo. CatG, uniquely among endocytic proteases tested, initiated cleavage of detergent-solubilized native and recombinant soluble MHC II molecules. CatG cleaved human leukocyte antigen (HLA)-DR isolated from both HLA-DM-expressing and DM-null cells. Even following CatG cleavage, peptide binding was retained by pre-loaded, soluble recombinant HLA-DR. MHC II cleavage occurred on the loop between fx1 and fx2 of the membrane-proximal b2 domain. All allelic variants of HLA-DR tested and murine I-A g7 class II molecules were susceptible, whereas murine I-E k and HLA-DM were not, consistent with their altered sequence at the P1' position of the CatG cleavage site. CatG effects were reduced on HLA-DR molecules with DRB mutations in the region implicated in interaction with HLA-DM. In contrast, addition of CatG to intact B-lymphoblastoid cell lines (B-LCLs) did not cause degradation of membrane-bound MHC II. Moreover, inhibition or genetic ablation of CatG in primary antigen-presenting cells did not cause accumulation of MHC II molecules. Thus, in vivo, the CatG cleavage site is sterically inaccessible or masked by associated molecules. A combination of intrinsic and context-dependent proteolytic resistance may allow peptide capture by MHC II molecules in harshly proteolytic endocytic compartments, as well as persistent antigen presentation in acute inflammatory settings with extracellular proteolysis.
Proceedings of the National Academy of Sciences, 2003
MHC class II-restricted antigen presentation plays a central role in the immune response against exogenous antigens. The association of invariant (Ii) chain with MHC class II dimers is required for proper antigen presentation to CD4 ؉ T cells by antigen-presenting cells. MHC class II complexes first traffic through the endocytic pathway to allow Ii chain degradation and antigenic peptide loading before their arrival at the cell surface. In recent years, a considerable effort has been directed toward the identification of proteases responsible for Ii chain degradation. Targeted gene deletion in mice has allowed a precise description of the cysteine proteases involved in the last step of Ii chain degradation. By using nonspecialized cellular models expressing MHC II molecules, we are now exploring the contribution of known cysteine proteases to human Ii chain processing. Surprisingly and contrary to the situation in mouse, cathepsin S was found to be the only human cysteine protease able to efficiently degrade the Ii-p10 fragment in epithelial cells. This selectivity has implications for thymic selection and indicates that differences between man and mice are probably more profound at this level than expected.
Quantifying Cathepsin S Activity in Antigen Presenting Cells Using a Novel Specific Substrate
Journal of Biological Chemistry, 2008
Cathepsin S (CatS) is a lysosomal cysteine protease belonging to the papain superfamily. Because of the relatively broad substrate specificity of this family, a specific substrate for CatS is not yet known. Based on a detailed study of the CatS endopeptidase specificity, using six series of internally quenched fluorescent peptides, we were able to design a specific substrate for CatS. The peptide series was based on the sequence GRWHTVGL-RWE-Lys(Dnp)-DArg-NH 2 , which shows only one single cleavage site between Gly and Leu and where every substrate position between P-3 and P-3 was substituted with up to 15 different amino acids. The endopeptidase specificity of CatS was mainly determined by the P-2, P-1, and the P-3 substrate positions.
FEBS Letters, 1993
The complex of cathepsin L and the fragment of the MHC class II-associated invariant chain was purified from human kidney. M, of the complex, as determined by gel filtration, is about 40,000. Both components were identified by amino acid and sequence analyses. The bound invariant chain fragment is almost identical to the additional segment found in p41, but not in the p3 1 form of the invariant chain. The complex has significantly enhanced stability at neutral and slightly alkaline pH, and reduced proteolytic activity against the synthetic substrate Z-Phe-Arg-MCA compared to free cathepsin L. The complex exhibits no enzymatic activity against the protein substrate azocasein. For the first time, the invariant chain was found in a complex with a protein, which was not an MHC molecule. Cathepsin L; Cysteine proteinase; MHC class II molecule; Invariant chain; p41