Browsing by Subject "antigen processing"
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- Research DataAffinity-dependent peptide loading drives sequential disassembly of the peptide-loading complex2026-07-21Antigen presentation by major histocompatibility complex class I (MHC I) depends on the accurate assembly of peptide–MHC I complexes, yet the molecular mechanism coupling peptide loading to peptide-loading complex (PLC) disassembly remains unresolved. Progress has been limited by the lack of an in vitro system that faithfully recapitulates the native membrane environment. Here, we reconstitute intact human PLCs in liposomes, enabling analysis of peptide loading, MHC I release, and post-release glycan processing under membrane-reconstituted conditions. We show that high-affinity peptide binding triggers allomorph-specific release of MHC I from the PLC, independent of the conformational state of the transporter associated with antigen processing (TAP). Furthermore, glucosidase II-mediated N-glycan trimming and calreticulin dissociation occur predominantly after MHC I exits the PLC, revealing a sequential disassembly mechanism. These findings define the molecular sequence of events underlying PLC disassembly and provide a versatile platform for mechanistic studies of antigen processing.
7 3 - Research DataQuantitative nanoscale imaging shows peptide–MHC I complexes are monomeric and spatially regulated in human dendritic cells2026-07-13Jacobs, OliviaMenche, TanjaHöper, CindyGerhards, FrédéricFucek, IvicaVascotto, FulviaDietz, Marina S.Heilemann, MikeMajor histocompatibility complex class I (MHC I) molecules present antigenic peptides to cytotoxic T cells, a process central to immune surveillance. However, the nanoscale spatial organization of peptide–MHC I (pMHC I) on human dendritic cells (DCs), key initiators of cytotoxic T cell responses, remains largely unexplored. Here, we combine high-affinity soluble T cell receptors with DNA-based point accumulation for imaging in nanoscale topography (DNA-PAINT) to quantitatively map and count defined pMHC I complexes at single-molecule resolution on HLA-A*02:01-expressing cells and primary human monocyte-derived DCs. We found no evidence for higher-order pMHC I nanoclusters under conditions of extracellular peptide exchange or physiological intracellular loading. Instead, detected signals correspond to individual pMHC I complexes. Notably, DC differentiation and activation modulate pMHC I surface abundance and spatial compartmentalization. These findings refine current models of antigen presentation by emphasizing regulation through surface density and spatial distribution, and establish a quantitative framework for epitope-specific, single-molecule quantification of antigen presentation in human immune cells.
7 16 - Research DataReconstitution of glycan-driven MHC I recycling reveals calreticulin as mediator between TAPBPR and tapasin2026-05-22Müller, Ines K.Popovic, NikoMorgner, NinaTrowitzsch, SimonProtein folding in the endoplasmic reticulum (ER) relies on N-linked glycosylation and glycan remodeling to guide quality control. Major histocompatibility complex class I (MHC I) molecules, essential for adaptive immunity, undergo a specialized maturation pathway involving the peptide-loading complex (PLC), the editor TAPBPR, the UDP-glucose:glycoprotein glucosyltransferase, and the lectin chaperone calreticulin. However, how glycan-dependent mechanisms coordinate MHC I transfer between these factors has remained unclear. Using a fully reconstituted system, we show that retrograde transfer of peptide-receptive MHC I from TAPBPR to tapasin requires calreticulin recognition of monoglucosylated MHC I glycans. While calreticulin’s C-terminal acidic helix is dispensable for releasing reglucosylated MHC I from TAPBPR, it is essential for productive docking of MHC I onto tapasin. These findings reveal a glycan-surveillance mechanism that enables retrieval of suboptimally loaded MHC I molecules missed by the initial quality control at the PLC. Our work defines a glycan-dependent chaperone network, finely tuned by a combination of low-micromolar interactions between the constituents, that ensures efficient MHC I maturation and illustrates fundamental principles of ER protein quality control.
3 19 - Research DataReconstitution of glycan-driven MHC I recycling reveals calreticulin as mediator between TAPBPR and tapasin2025-04-24Popovic, NikoRath, TobiasMorgner, NinaTrowitzsch, SimonTampé, RobertHeinke, Tim Julius (DataCollector)Fahim, Amin (DataCollector)Trowitzsch, Simon (ContactPerson)Tampé, Robert (ContactPerson)Popovic, Niko (DataCollector)Rath, Tobias (DataCollector)Morgner, Nina (ContactPerson)Protein folding in the endoplasmic reticulum (ER) is essential for about one-third of the mammalian proteome. N-linked glycosylation and subsequent glycan remodeling barcodes glycoproteins during their maturation in the ER. Major histocompatibility complex class I (MHC I) molecules, key for adaptive immunity, rely on a dedicated quality control cycle that involves specialized chaperones and glycan-modifying enzymes for their maturation and loading of immunogenic peptides. However, the functional interplay of the MHC I editors tapasin as part of the peptide-loading complex (PLC), TAP-binding protein-related (TAPBPR), the UDP-glucose:glycoprotein glucosyltransferase 1 (UGGT1), and calreticulin in glycan-dependent transfer of MHC I clients has not been determined in a reconstituted system. With isolated components, we show that transfer of peptide-receptive MHC I from the downstream quality control factor TAPBPR back to tapasin depends on the recognition of the monoglucosylated glycan of MHC I by calreticulin. While calreticulin’s C-terminal acidic helix is dispensable for disengaging reglucosylated MHC I from TAPBPR, it is essential for docking MHC I onto tapasin. Our data provide a mechanistic basis for glycan-surveillance by calreticulin necessary for retrograde trafficking of misfolded or suboptimally loaded MHC I that escaped the first quality control at the PLC and were trapped by TAPBPR. Such finetuned dynamic network of glycan-dependent and MHC I-specific chaperones guarantees maturation of MHC I molecules and highlight the fundamental processes driving ER protein quality control.
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