Abstract
dc:description.abstractCytotoxic T-lymphocytes must recognise and respond in a highly antigen specific manner to tackle pathological threats, such as virus-infected or malignant cells. This is aided by cell surface glycoproteins called the major histocompatibility complex (MHC). MHC class I (MHC-I) molecules are polymorphic structures that can display a large repertoire of peptides. Peptide loading occurs in the context of a multi-protein machinery called the peptide loading complex (PLC). A key player in the initial loading of peptides on MHC-I is owed to the peptide editor, tapasin. More recently, a second peptide editor, named TAPBPR, was discovered. Together, they shape the MHC-I presented peptidome. However, unlike its homologue tapasin, TAPBPR catalyses peptide exchange independently of the PLC. It works closely with UDP-glucose: glycoprotein glucosyltransferase 1 (UGT1) to filter sub-optimally loaded MHC-I molecules before export to the cell surface. To date, extensive insight on TAPBPR’s biology has been gained using cell-free assays, structural studies and via its characterisation in the cervical cancer cell line, HeLa. Currently, there is limited data on TAPBPR in other cell types. Through this body of work, TAPBPR has been characterised in peripheral blood mononuclear cells (PBMCs). TAPBPR protein was found to be expressed in PBMCs, with the most abundance in monocytes. Furthermore, it became clear that TAPBPR exhibits donor-to-donor variability and could potentially be expressed as alternative spliced products. Historically, mass spectrometry led to the discovery of TAPBPR’s interaction with MHC-I and UGT1 in HeLa cells. However, in this project, by using a more physiologically relevant model system, numerous novel putative binding partners in cell lines (B-lymphocytes and monocytes) and primary immune cells were identified. Intriguingly, interactomic studies revealed the association of TAPBPR with non-classical MHC-I molecules. Taking specific interest in HLA-F, an enigmatic MHC-I molecule, the association was confirmed through immunoprecipitation, followed by western blotting, and also through the reciprocal immunoprecipitation, in B-lymphocytes. The results indicated that TAPBPR is likely interacting with HLA-F bound to β2-microglobulin (β2m). To unravel the biology behind this interaction, phenotyping was performed. HLA-F is thought to be recognised by leukocyte immunoglobulin-like receptors (LILRs) or killer-cell immunoglobulin-like receptors (KIRs), and only recently deemed as capable of presenting peptides. These studies used soluble HLA-F, when in fact HLA-F naturally has a transmembrane domain. Immunopeptidomic studies performed here, from HLA-F pull downs, revealed it indeed presents peptides in a non-conventional manner. HLA-F/β2m heterodimers accommodate exceptionally long peptides, as is the case for MHC class II peptides, with a striking preference for charged anchor residues at the C-termini. Cumulatively, these findings provide new understanding into TAPBPR’s role in antigen processing and presentation in the context of non-classical MHC-I.
Degree
thesis:*- Name dc:type.qualificationname
- Doctor of Philosophy (PhD)
- Level dc:type.qualificationlevel
- Doctoral
- Grantor dc:publisher.institution
- University of Cambridge
- Year dc:date.issued
- 2023
Author and committee
dc:creator, dc:contributor.*- Author dc:creator
-
- Satti, Reem
- Advisor dc:contributor.advisor
-
- Boyle, Louise
Subjects
dc:subject × 3Rights
dc:rightsIdentifiers
dc:identifier.*- DOI dc:identifier.doi
- https://doi.org/10.17863/CAM.97078
- OAI identifier oai:identifier
- oai:www.repository.cam.ac.uk:1810/350698