Endocytosis targets exogenous material selectively to cathepsin S in live human dendritic cells, while cell-penetrating peptides mediate nonselective transport to cysteine cathepsins.
The way the MHC II-associated proteolytic system of APC handles exogenous antigen is key to the stimulation of the T cell in infections and immunotherapy settings. Using a cell-impermeable, activity-based probe (ABP) for papain cathepsins, the most abundant type of endocytic proteases, we have simulated the encounter between exogenous antigen and endocytic proteases in live human monocyte-derived dendritic cells (MO-DC). Although cathepsin S (CatS), -B, -H, and -X were active in DC-derived endocytic fractions in vitro, the peptide-size tracer was routed selectively to active CatS after internalization by macropinocytosis. Blocking of the vacuolar adenosine triphosphatase abolished this CatS-selective targeting, and LPS-induced maturation of DC resulted in degradation of active CatS. Conjugation of the ABP to a protein facilitated the delivery to endocytic proteases and resulted in labeling of sizable amounts of CatB and CatX, although CatS still remained the major protease reached by this construct. Conjugation of the probe to a cell-penetrating peptide (CPP) routed the tracer to the entire panel of intracellular cathepsins, independently from endocytosis or LPS stimulation. Thus, different means of internalization result in differential targeting of active cathepsins in live MO-DC. CPP may serve as vehicles to target antigen more efficiently to protease-containing endocytic compartments.
Reich
Michael
M
Department of Medicine II, Institute for Cell Biology, and Medical and Natural Sciences Research Center, University of Tübingen, Germany.
van Swieten
Paul F
PF
Sommandas
Vinod
V
Kraus
Marianne
M
Fischer
Rainer
R
Weber
Ekkehard
E
Kalbacher
Hubert
H
Overkleeft
Herman S
HS
Driessen
Christoph
C
D000251:Adenosine Triphosphatases / Q000378:metabolism / Q000502:physiology; D000818:Animals; D001692:Biological Transport; D002403:Cathepsins / Q000378:metabolism*; D002478:Cells, Cultured; D003546:Cysteine Endopeptidases / Q000378:metabolism; D003713:Dendritic Cells / Q000378:metabolism*; D004705:Endocytosis; D000949:Histocompatibility Antigens Class II / Q000378:metabolism; D006801:Humans; D007930:Leucine / Q000031:analogs & derivatives* / Q000493:pharmacokinetics; D051379:Mice; D008807:Mice, Inbred BALB C; D008863:Microspheres; D015335:Molecular Probes / Q000378:metabolism; D015394:Molecular Structure; D009000:Monocytes / Q000378:metabolism / Q000502:physiology; D010455:Peptides / Q000493:pharmacokinetics; D010587:Phagocytosis; D021381:Protein Transport; D019809:Streptavidin / Q000493:pharmacokinetics; D014617:Vacuoles / Q000201:enzymology
C419228:DCG 04; D000949:Histocompatibility Antigens Class II; D015335:Molecular Probes; D010455:Peptides; D019809:Streptavidin; D002403:Cathepsins; D003546:Cysteine Endopeptidases; C028217:cathepsin S; D000251:Adenosine Triphosphatases; D007930:Leucine