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

作者：Anmole Gursev; Kuang Xiaomei T; Toyoda Mako; Martin Eric; Shahid Aniqa; Le Anh Q; Markle Tristan; Baraki Bemuluyigza; Jones R Brad; Ostrowski Mario A; Ueno Takamasa; Brumme Zabrina L; Brockman Mark A^*

来源：Journal of Immunological Methods, 2015, 426: 104-113.

DOI：10.1016/j.jim.2015.08.010

• Human immunodeficiency virus
• Nef
• HLA class I
• Immune evasion
• T cell receptor

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.

• 出版日期2015-11