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Presentation of exogenous antigens on major histocompatibility complex (MHC) class I and MHC class II molecules is differentially regulated during dendritic cell maturation.

Delamarre L, Holcombe H, Mellman I - J. Exp. Med. (2003)

Bottom Line: Unlike MHC II, these events do not involve a marked redistribution of preexisting MHC I molecules from intracellular compartments to the DC surface.In contrast, formation of peptide-MHC I complexes from endogenous cytosolic antigens occurs even in unstimulated, immature DCs.Thus, the MHC I and MHC II pathways of antigen presentation are differentially regulated during DC maturation.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Section of Immunobiology, Ludwig Institute for Cancer Research, Yale University School of Medicine, New Haven, CT 06520-8002, USA.

ABSTRACT
During maturation, dendritic cells (DCs) regulate their capacity to process and present major histocompatibility complex (MHC) II-restricted antigens. Here we show that presentation of exogenous antigens by MHC I is also subject to developmental control, but in a fashion strikingly distinct from MHC II. Immature mouse bone marrow-derived DCs internalize soluble ovalbumin and sequester the antigen intracellularly until they receive an appropriate signal that induces cross presentation. At that time, peptides are generated in a proteasome-dependent fashion and used to form peptide-MHC I complexes that appear at the plasma membrane. Unlike MHC II, these events do not involve a marked redistribution of preexisting MHC I molecules from intracellular compartments to the DC surface. Moreover, out of nine stimuli well known to induce the phenotypic maturation of DCs and to promote MHC II presentation, only two (CD40 ligation, disruption of cell-cell contacts) activated cross presentation on MHC I. In contrast, formation of peptide-MHC I complexes from endogenous cytosolic antigens occurs even in unstimulated, immature DCs. Thus, the MHC I and MHC II pathways of antigen presentation are differentially regulated during DC maturation.

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Cross presentation of OVA internalized prior DC activation. (A) Immature B6D2F1 DC were pulsed with OVA or BSA as a control (1 mg/ml) for 2 h, washed, and chased for 0 to 48 h before activation for 7 h with LPS and cluster disruption. Cells were then fixed and added to OT.1 T cells. (B) After a 24 h chase and 30 min before activation, epoxomicin 1 μM final was added to the cells and was also present during the 7 h stimulation period. As a control we used DMSO in which the drug was stocked. Cells were then fixed and added to OT.1 T cells. Right panel: OVA-pulsed cells and T cells were cultured in presence of OVA peptide. At 24 h, as marker of T cell activation IL-2 secretion was measured. One representative experiment out of three is shown, and the values represent the mean of triplicate wells. (*) indicates below level of detection.
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fig6: Cross presentation of OVA internalized prior DC activation. (A) Immature B6D2F1 DC were pulsed with OVA or BSA as a control (1 mg/ml) for 2 h, washed, and chased for 0 to 48 h before activation for 7 h with LPS and cluster disruption. Cells were then fixed and added to OT.1 T cells. (B) After a 24 h chase and 30 min before activation, epoxomicin 1 μM final was added to the cells and was also present during the 7 h stimulation period. As a control we used DMSO in which the drug was stocked. Cells were then fixed and added to OT.1 T cells. Right panel: OVA-pulsed cells and T cells were cultured in presence of OVA peptide. At 24 h, as marker of T cell activation IL-2 secretion was measured. One representative experiment out of three is shown, and the values represent the mean of triplicate wells. (*) indicates below level of detection.

Mentions: Internalized antigens can be sequestered by immature DCs for extended periods (at least 60 h) before being processed and loaded onto MHC II upon maturation (11). We next determined if immature DCs could also sequester antigens for cross presentation. OVA-pulsed DCs were chased for up to 48 h without further stimulation. At various times of chase, the DCs were “cluster disrupted,” and then incubated in presence of LPS for an additional 7 h before fixation and assay using OT.1 T cells. The DCs were capable of cross presenting OVA that had been internalized at least up to 48 h before receiving the activation stimulus (Fig. 6 A). While the amount of cross presentation decreased with time, at 24 and 48 h, significant OT.1 T cell responses were observed corresponding respectively to 45 and 25% of the amount of presentation exhibited by DCs activated and assayed immediately after exposure to OVA.


Presentation of exogenous antigens on major histocompatibility complex (MHC) class I and MHC class II molecules is differentially regulated during dendritic cell maturation.

Delamarre L, Holcombe H, Mellman I - J. Exp. Med. (2003)

Cross presentation of OVA internalized prior DC activation. (A) Immature B6D2F1 DC were pulsed with OVA or BSA as a control (1 mg/ml) for 2 h, washed, and chased for 0 to 48 h before activation for 7 h with LPS and cluster disruption. Cells were then fixed and added to OT.1 T cells. (B) After a 24 h chase and 30 min before activation, epoxomicin 1 μM final was added to the cells and was also present during the 7 h stimulation period. As a control we used DMSO in which the drug was stocked. Cells were then fixed and added to OT.1 T cells. Right panel: OVA-pulsed cells and T cells were cultured in presence of OVA peptide. At 24 h, as marker of T cell activation IL-2 secretion was measured. One representative experiment out of three is shown, and the values represent the mean of triplicate wells. (*) indicates below level of detection.
© Copyright Policy
Related In: Results  -  Collection

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fig6: Cross presentation of OVA internalized prior DC activation. (A) Immature B6D2F1 DC were pulsed with OVA or BSA as a control (1 mg/ml) for 2 h, washed, and chased for 0 to 48 h before activation for 7 h with LPS and cluster disruption. Cells were then fixed and added to OT.1 T cells. (B) After a 24 h chase and 30 min before activation, epoxomicin 1 μM final was added to the cells and was also present during the 7 h stimulation period. As a control we used DMSO in which the drug was stocked. Cells were then fixed and added to OT.1 T cells. Right panel: OVA-pulsed cells and T cells were cultured in presence of OVA peptide. At 24 h, as marker of T cell activation IL-2 secretion was measured. One representative experiment out of three is shown, and the values represent the mean of triplicate wells. (*) indicates below level of detection.
Mentions: Internalized antigens can be sequestered by immature DCs for extended periods (at least 60 h) before being processed and loaded onto MHC II upon maturation (11). We next determined if immature DCs could also sequester antigens for cross presentation. OVA-pulsed DCs were chased for up to 48 h without further stimulation. At various times of chase, the DCs were “cluster disrupted,” and then incubated in presence of LPS for an additional 7 h before fixation and assay using OT.1 T cells. The DCs were capable of cross presenting OVA that had been internalized at least up to 48 h before receiving the activation stimulus (Fig. 6 A). While the amount of cross presentation decreased with time, at 24 and 48 h, significant OT.1 T cell responses were observed corresponding respectively to 45 and 25% of the amount of presentation exhibited by DCs activated and assayed immediately after exposure to OVA.

Bottom Line: Unlike MHC II, these events do not involve a marked redistribution of preexisting MHC I molecules from intracellular compartments to the DC surface.In contrast, formation of peptide-MHC I complexes from endogenous cytosolic antigens occurs even in unstimulated, immature DCs.Thus, the MHC I and MHC II pathways of antigen presentation are differentially regulated during DC maturation.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Section of Immunobiology, Ludwig Institute for Cancer Research, Yale University School of Medicine, New Haven, CT 06520-8002, USA.

ABSTRACT
During maturation, dendritic cells (DCs) regulate their capacity to process and present major histocompatibility complex (MHC) II-restricted antigens. Here we show that presentation of exogenous antigens by MHC I is also subject to developmental control, but in a fashion strikingly distinct from MHC II. Immature mouse bone marrow-derived DCs internalize soluble ovalbumin and sequester the antigen intracellularly until they receive an appropriate signal that induces cross presentation. At that time, peptides are generated in a proteasome-dependent fashion and used to form peptide-MHC I complexes that appear at the plasma membrane. Unlike MHC II, these events do not involve a marked redistribution of preexisting MHC I molecules from intracellular compartments to the DC surface. Moreover, out of nine stimuli well known to induce the phenotypic maturation of DCs and to promote MHC II presentation, only two (CD40 ligation, disruption of cell-cell contacts) activated cross presentation on MHC I. In contrast, formation of peptide-MHC I complexes from endogenous cytosolic antigens occurs even in unstimulated, immature DCs. Thus, the MHC I and MHC II pathways of antigen presentation are differentially regulated during DC maturation.

Show MeSH