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Degradation of mouse invariant chain: roles of cathepsins S and D and the influence of major histocompatibility complex polymorphism.

Villadangos JA, Riese RJ, Peters C, Chapman HA, Ploegh HL - J. Exp. Med. (1997)

Bottom Line: Antigen-presenting cells (APC) degrade endocytosed antigens into peptides that are bound and presented to T cells by major histocompatibility complex (MHC) class II molecules.Class II molecules are delivered to endocytic compartments by the class II accessory molecule invariant chain (Ii), which itself must be eliminated to allow peptide binding.These observations suggest that, first, class II molecules associated with larger Ii remnants can be converted efficiently to class II-peptide complexes and, second, that most class II-associated peptides can still be generated in cells treated with inhibitors of cysteine proteases.

View Article: PubMed Central - PubMed

Affiliation: Center for Cancer Research, Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.

ABSTRACT
Antigen-presenting cells (APC) degrade endocytosed antigens into peptides that are bound and presented to T cells by major histocompatibility complex (MHC) class II molecules. Class II molecules are delivered to endocytic compartments by the class II accessory molecule invariant chain (Ii), which itself must be eliminated to allow peptide binding. The cellular location of Ii degradation, as well as the enzymology of this event, are important in determining the sets of antigenic peptides that will bind to class II molecules. Here, we show that the cysteine protease cathepsin S acts in a concerted fashion with other cysteine and noncysteine proteases to degrade mouse Ii in a stepwise fashion. Inactivation of cysteine proteases results in incomplete degradation of Ii, but the extent to which peptide loading is blocked by such treatment varies widely among MHC class II allelic products. These observations suggest that, first, class II molecules associated with larger Ii remnants can be converted efficiently to class II-peptide complexes and, second, that most class II-associated peptides can still be generated in cells treated with inhibitors of cysteine proteases. Surprisingly, maturation of MHC class II in mice deficient in cathepsin D is unaffected, showing that this major aspartyl protease is not involved in degradation of Ii or in generation of the bulk of antigenic peptides.

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The effect of cysteine protease inhibition on maturation of  MHC class II molecules varies widely among allelic products. (A) Spleen  cells of mice of the haplotypes indicated were pulse-labeled and chased for  4 h in the absence or presence of 1 mM leupeptin or 3 nM LHVS, and  their I-A molecules immunoprecipitated with an anti-I-Aα rabbit serum  (44). Immunoprecipitates were run on 12.5% SDS-PAGE without (top  half) or after (bottom half) boiling. (B) Amount of I-A SDS-stable dimers  generated in leupeptin-treated splenocytes of different haplotypes relative  to their control counterparts. The amount of SDS-stable I-Ad complexes  in control cells was too small to perform a reliable comparison to the  drug-treated samples. (C) Same as in B, for the LHVS-treated samples.  (D) Cbz–[125I]–Tyr–Ala–CN2 labeling of H-2d, H-2b, and H-2k splenocytes.
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Figure 4: The effect of cysteine protease inhibition on maturation of MHC class II molecules varies widely among allelic products. (A) Spleen cells of mice of the haplotypes indicated were pulse-labeled and chased for 4 h in the absence or presence of 1 mM leupeptin or 3 nM LHVS, and their I-A molecules immunoprecipitated with an anti-I-Aα rabbit serum (44). Immunoprecipitates were run on 12.5% SDS-PAGE without (top half) or after (bottom half) boiling. (B) Amount of I-A SDS-stable dimers generated in leupeptin-treated splenocytes of different haplotypes relative to their control counterparts. The amount of SDS-stable I-Ad complexes in control cells was too small to perform a reliable comparison to the drug-treated samples. (C) Same as in B, for the LHVS-treated samples. (D) Cbz–[125I]–Tyr–Ala–CN2 labeling of H-2d, H-2b, and H-2k splenocytes.

Mentions: Splenocytes of mice of the d, b, k, s, and u haplotypes were pulse labeled for 30 min and chased for 4 h in the absence or presence of leupeptin or LHVS. I-A molecules were then immunoprecipitated with a rabbit serum that reacts specifically with the COOH-terminal segment of the I-Aα chain (44) (Fig. 4 A). The amount of the I-Aα subunit in the pulsed samples was larger than in their chased counterparts. This is probably due to the presence of excess free α chains in the pulsed samples that fail to assemble with β subunits and are degraded during the chase. Because the anti-I-Aα serum can recognize free α chains (44; see Fig 1 B), these can be recovered along with the αβ dimers.


Degradation of mouse invariant chain: roles of cathepsins S and D and the influence of major histocompatibility complex polymorphism.

Villadangos JA, Riese RJ, Peters C, Chapman HA, Ploegh HL - J. Exp. Med. (1997)

The effect of cysteine protease inhibition on maturation of  MHC class II molecules varies widely among allelic products. (A) Spleen  cells of mice of the haplotypes indicated were pulse-labeled and chased for  4 h in the absence or presence of 1 mM leupeptin or 3 nM LHVS, and  their I-A molecules immunoprecipitated with an anti-I-Aα rabbit serum  (44). Immunoprecipitates were run on 12.5% SDS-PAGE without (top  half) or after (bottom half) boiling. (B) Amount of I-A SDS-stable dimers  generated in leupeptin-treated splenocytes of different haplotypes relative  to their control counterparts. The amount of SDS-stable I-Ad complexes  in control cells was too small to perform a reliable comparison to the  drug-treated samples. (C) Same as in B, for the LHVS-treated samples.  (D) Cbz–[125I]–Tyr–Ala–CN2 labeling of H-2d, H-2b, and H-2k splenocytes.
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Figure 4: The effect of cysteine protease inhibition on maturation of MHC class II molecules varies widely among allelic products. (A) Spleen cells of mice of the haplotypes indicated were pulse-labeled and chased for 4 h in the absence or presence of 1 mM leupeptin or 3 nM LHVS, and their I-A molecules immunoprecipitated with an anti-I-Aα rabbit serum (44). Immunoprecipitates were run on 12.5% SDS-PAGE without (top half) or after (bottom half) boiling. (B) Amount of I-A SDS-stable dimers generated in leupeptin-treated splenocytes of different haplotypes relative to their control counterparts. The amount of SDS-stable I-Ad complexes in control cells was too small to perform a reliable comparison to the drug-treated samples. (C) Same as in B, for the LHVS-treated samples. (D) Cbz–[125I]–Tyr–Ala–CN2 labeling of H-2d, H-2b, and H-2k splenocytes.
Mentions: Splenocytes of mice of the d, b, k, s, and u haplotypes were pulse labeled for 30 min and chased for 4 h in the absence or presence of leupeptin or LHVS. I-A molecules were then immunoprecipitated with a rabbit serum that reacts specifically with the COOH-terminal segment of the I-Aα chain (44) (Fig. 4 A). The amount of the I-Aα subunit in the pulsed samples was larger than in their chased counterparts. This is probably due to the presence of excess free α chains in the pulsed samples that fail to assemble with β subunits and are degraded during the chase. Because the anti-I-Aα serum can recognize free α chains (44; see Fig 1 B), these can be recovered along with the αβ dimers.

Bottom Line: Antigen-presenting cells (APC) degrade endocytosed antigens into peptides that are bound and presented to T cells by major histocompatibility complex (MHC) class II molecules.Class II molecules are delivered to endocytic compartments by the class II accessory molecule invariant chain (Ii), which itself must be eliminated to allow peptide binding.These observations suggest that, first, class II molecules associated with larger Ii remnants can be converted efficiently to class II-peptide complexes and, second, that most class II-associated peptides can still be generated in cells treated with inhibitors of cysteine proteases.

View Article: PubMed Central - PubMed

Affiliation: Center for Cancer Research, Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.

ABSTRACT
Antigen-presenting cells (APC) degrade endocytosed antigens into peptides that are bound and presented to T cells by major histocompatibility complex (MHC) class II molecules. Class II molecules are delivered to endocytic compartments by the class II accessory molecule invariant chain (Ii), which itself must be eliminated to allow peptide binding. The cellular location of Ii degradation, as well as the enzymology of this event, are important in determining the sets of antigenic peptides that will bind to class II molecules. Here, we show that the cysteine protease cathepsin S acts in a concerted fashion with other cysteine and noncysteine proteases to degrade mouse Ii in a stepwise fashion. Inactivation of cysteine proteases results in incomplete degradation of Ii, but the extent to which peptide loading is blocked by such treatment varies widely among MHC class II allelic products. These observations suggest that, first, class II molecules associated with larger Ii remnants can be converted efficiently to class II-peptide complexes and, second, that most class II-associated peptides can still be generated in cells treated with inhibitors of cysteine proteases. Surprisingly, maturation of MHC class II in mice deficient in cathepsin D is unaffected, showing that this major aspartyl protease is not involved in degradation of Ii or in generation of the bulk of antigenic peptides.

Show MeSH
Related in: MedlinePlus