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Content archived on 2024-05-29

The role of the intracellular peptidases in the immune system and in the cellular metabolism

Objective

MHC class I molecules present peptides from intracellular antigens to the immune system. The peptides are produced by the intracellular proteasome and further trimmed and destroyed by various unrelated peptidases. Only a fraction (0.01%) of these peptides escape peptidase destruction through import into the ER by the peptide transporter TAP before they can bind to resident MHC class I molecules for presentation at the plasma membrane. Since the peptidase activity plays an important role in the selection of peptides for presentation, we propose to study these using microinjection of internally quenched peptides and inhibitors, to measure peptidase activity in living cells.

I aim to: 1. Test with a large set of internally quenched peptides the subspecializati on of intracellular peptidases for substrate (peptide) length and sequence. 2. Identify the respective peptidases responsible for the detected subspecializations. Chemical inhibitors and RNAi for TOP, BH and TPPII have been defined and their effects on pep tide destruction will be measured. 3. Test the effect of the peptidases on the petpidome as presented by MHC class I molecules. Peptidases will be inhibited and the resulting peptidome will be assayed by comparative mass spectrometry using differently labelled amino acids. 4. Test whether different cells employ different peptidase activities. The Neefjes lab has indications that dendritic cells change their peptidase activity and I will study this and the effect of various activation stages of these cells in more detail. 5. Combine these findings with the results on manipulation for peptidase activity to determine the effect of peptidase activity on the outcome of MHC class I response in dendritic cells.

The experiments should reveal the role of cytosolic peptidase activities in antigen presentation by MHC class I molecules and add information to better predict the MHC class I peptidome on the basis of the primary sequence of the antigen.

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Keywords

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Topic(s)

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Call for proposal

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FP6-2005-MOBILITY-5
See other projects for this call

Funding Scheme

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EIF - Marie Curie actions-Intra-European Fellowships

Coordinator

HET NEDERLANDS KANKER INSTITUUT
EU contribution
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Total cost

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