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Processing of a multiple membrane spanning Epstein-Barr virus protein for CD8(+) T cell recognition reveals a
G Lautscham1, S Mayrhofer, G Taylor
1Cancer Research Campaign Institute for Cancer Studies and Medical Research Council Centre for Immune Regulation, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom.
Abstract:
Epstein-Barr virus (EBV) latent membrane protein (LMP)2 is a multiple membrane spanning molecule which lacks ectodomains projecting into the lumen of the endoplasmic reticulum (ER). Human CD8(+) cytotoxic T lymphocytes (CTL)s recognize a number of epitopes within LMP2. Assays with epitope-specific CTLs in two different cell backgrounds lacking the transporter associated with antigen processing (TAP) consistently show that some, but not all, LMP2 epitopes are presented in a TAP-independent manner. However, unlike published examples of TAP-independent processing from endogenously expressed antigens, presentation of TAP-independent LMP2 epitopes was abrogated by inhibition of proteasomal activity. We found a clear correlation between hydrophobicity of the LMP2 epitope sequence and TAP independence, and experiments with vaccinia minigene constructs expressing cytosolic epitope peptides confirmed that these more hydrophobic peptides were selectively able to access the HLA class I pathway in TAP-negative cells. Furthermore, the TAP-independent phenotype of particular epitope sequences did not require membrane location of the source antigen since (i) TAP-independent LMP2 epitopes inserted into an EBV nuclear antigen and (ii) hydrophobic epitope sequences native to EBV nuclear antigens were both presented in TAP-negative cells. We infer that there is a proteasome-dependent, TAP-independent pathway of antigen presentation which hydrophobic epitopes can selectively access.
Insights
Certain Epstein-Barr virus (EBV) latent membrane protein 2 (LMP2) epitopes are presented independently of the transporter associated with antigen processing (TAP). This pathway relies on proteasomal activity and favors hydrophobic epitope sequences.
Area of Science:
- Immunology
- Virology
- Molecular Biology
Background:
- Epstein-Barr virus (EBV) latent membrane protein 2 (LMP2) is a transmembrane protein crucial in EBV pathogenesis.
- Human CD8+ cytotoxic T lymphocytes (CTLs) recognize viral epitopes presented by HLA class I molecules.
- Antigen processing and presentation typically involve the transporter associated with antigen processing (TAP).
Purpose of the Study:
- To investigate the antigen processing and presentation pathways of Epstein-Barr virus (EBV) latent membrane protein 2 (LMP2) epitopes.
- To determine the role of the transporter associated with antigen processing (TAP) in presenting LMP2-derived epitopes.
- To elucidate the characteristics of LMP2 epitopes presented in a TAP-independent manner.
Main Methods:
- Utilized epitope-specific cytotoxic T lymphocytes (CTLs) in transporter associated with antigen processing (TAP)-deficient cell lines.
- Assessed the impact of proteasomal inhibition on epitope presentation.
- Employed vaccinia minigene constructs to express epitope peptides.
- Investigated epitope presentation from both membrane-bound and non-membrane-bound EBV antigens.
Main Results:
- Some Epstein-Barr virus (EBV) latent membrane protein 2 (LMP2) epitopes were presented in a transporter associated with antigen processing (TAP)-independent manner.
- TAP-independent presentation of LMP2 epitopes was dependent on proteasomal activity.
- A strong correlation was observed between epitope sequence hydrophobicity and TAP-independent presentation.
- Hydrophobic epitopes accessed the HLA class I pathway in TAP-negative cells, irrespective of their source antigen's membrane localization.
Conclusions:
- A proteasome-dependent, TAP-independent pathway for antigen presentation exists.
- This pathway selectively accommodates hydrophobic epitopes.
- Findings challenge conventional models of antigen processing and presentation for viral antigens.