A robust and scalable TCR-based reporter cell assay to measure HIV-1 Nef-mediated T cell immune evasion

Journal of Immunological Methods
Gursev AnmoleMark A Brockman

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, o...Continue Reading

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Citations

Oct 4, 2017·The Journal of Immunology : Official Journal of the American Association of Immunologists·Chike O AbanaSimon A Mallal
Sep 12, 2015·Journal of Virology·Aniqa ShahidZabrina L Brumme
Nov 28, 2018·Nature Communications·Funsho OgunsholaZaza M Ndhlovu
Jan 17, 2020·Journal of Medical Virology·Jaclyn K MannThumbi Ndung'u
Sep 10, 2020·Proceedings of the National Academy of Sciences of the United States of America·Mark M PainterKathleen L Collins
Sep 3, 2020·The Journal of Biological Chemistry·Ryan P Staudt, Thomas E Smithgall
Nov 7, 2019·Cell Reports·Steven W JinMark A Brockman

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