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A robust and scalable TCR-based reporter cell assay to measure HIV-1 Nef-mediated T cell immune evasion
- Source :
- Journal of Immunological Methods. 426:104-113
- Publication Year :
- 2015
- Publisher :
- Elsevier BV, 2015.
-
Abstract
- HIV-1 evades cytotoxic T cell responses through Nef-mediated downregulation of HLA class I molecules from the infected cell surface. Methods to quantify the impact of Nef on T cell recognition typically employ patient-derived T cell clones; however, these assays are limited by the cost and effort required to isolate and maintain primary cell lines. The variable activity of different T cell clones and the limited number of cells generated by re-stimulation can also hinder assay reproducibility and scalability. Here, we describe a heterologous T cell receptor reporter assay and use it to study immune evasion by Nef. Induction of NFAT-driven luciferase following co-culture with peptide-pulsed or virus-infected target cells serves as a rapid, quantitative and antigen-specific measure of T cell recognition of its cognate peptide/HLA complex. We demonstrate that Nef-mediated downregulation of HLA on target cells correlates inversely with T cell receptor-dependent luminescent signal generated by effector cells. This method provides a robust, flexible and scalable platform that is suitable for studies to measure Nef function in the context of different viral peptide/HLA antigens, to assess the function of patient-derived Nef alleles, or to screen small molecule libraries to identify novel Nef inhibitors.
- Subjects :
- T cell
Immunology
Receptors, Antigen, T-Cell
Down-Regulation
Human leukocyte antigen
Streptamer
Biology
gag Gene Products, Human Immunodeficiency Virus
Jurkat Cells
Immune system
medicine
Humans
Immunology and Allergy
Cytotoxic T cell
nef Gene Products, Human Immunodeficiency Virus
HLA Complex
Immune Evasion
Immunoassay
Reporter gene
NFATC Transcription Factors
Histocompatibility Antigens Class I
T-cell receptor
Molecular biology
Coculture Techniques
medicine.anatomical_structure
HIV-1
T-Lymphocytes, Cytotoxic
Subjects
Details
- ISSN :
- 00221759
- Volume :
- 426
- Database :
- OpenAIRE
- Journal :
- Journal of Immunological Methods
- Accession number :
- edsair.doi.dedup.....8cd4922ac4cf7296b929c81e509dea81