A glycan-dependent chaperone network is defined, 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.
Abstract
Protein 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.
An overview of the PLC as a major determinant of tumor immune escape and a potential therapy target is provided and existing or evolving therapeutic approaches to recover/reprogram PLC functions are explored.
O. Allela, Abdulkareem Shareef, Hayder Naji Sameer et al.· Current Medical Science· 0 citations
Abstract Class I MHC peptide (MHC-Ip) multimers are well-established reagents that detect antigen-specific T cells. The classical method for production of MHC-Ip multimers begins with the expression of MHC heavy chains (HCs) and β2-microglobulin (β2m) subunits as inclusion bodies in Escherichia coli and is followed by denaturant solubilization, in vitro folding in the presence of a defined peptide ligand, and purification by size exclusion chromatography. This protocol is labor intensive, difficult to scale, and represents a significant bottleneck in application of the technology. Herein, we present a novel method for the expression in eukaryotic cells of secreted peptide exchange–competent class I MHC proteins in their native conformation. In this method, expression constructs are engineered as bimolecular complexes composed of an MHC HC and a β2m molecule covalently linked at its amino terminus to an MHC-binding peptide through a flexible peptide linker containing a defined protease site. Upon proteolysis, the original peptide occupant of the MHC binding site dissociates and is easily replaced with a synthetic peptide. When leucine zippers are added to the carboxyl terminus of each subunit, protease cleavage of the linker results in a stable HC/β2m complex that can be isolated and stored for subsequent peptide loading. Using this method, we have produced homogeneous MHC-Ip complexes for 25 class I MHC alleles and demonstrated that tetramers produced in this way are equivalent to conventionally produced tetramers for T-cell staining.
V. Ramachandiran, J. Shires, Richard Willis et al.· Journal of Immunology· 0 citations
A unified structure-energy-dynamics model explaining how Post-translational modifications function as atomic-level chemical switches in antigen presentation is established, establishing a unified structure-energy-dynamics model explaining how PTMs function as atomic-level chemical switches in antigen presentation.
The main mechanisms of antigen acquisition, endocytic/lysosomal factors controlling MHCII-restricted processing and presentation, and evidence linking lysosomal dysfunction to autoimmunity are explored.
Gabriele Sergio Colangelo, Kyra J Cowan, Federico Riccardi Sirtori et al.· Journal of Immunology· 0 citations
This review describes the development of pathway-centered mechanistic synthesis across DNA and RNA virus families and further integrate innate immune antagonism, endoplasmic reticulum stress, antigen-presentation competence, cross-presentation limits, and virus-shaped peptide landscapes into a unified framework for understanding viral control of MHC-I output and its translational implications.
Yu Ye, Ying Zhang, Haobing Nie et al.· FEMS Microbiology Reviews· 0 citations
Hydrophilic interaction chromatography is utilized to enrich glycopeptides from both peptides bound to HLA and digest of HLA proteins that obtained from immunoprecipitation of MHC complex from cell lysate that enables the characterization of glycosylation of MHC by database search.
Rui Chen· Methods in molecular biology· 0 citations
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