Impact of conditional autophagy knockout on cytotoxic CD8 T cells at day 7 and day 8 after influenza infection. Mus musculus

To investigate the role of autophagy on cytotoxic CD8 T cells, we developed a new mouse that expresses an Atg5 gene with loxP sites up- and downstream of the exon 3, Cre-recombinase, T cell receptor specific for the OVA peptide SIINFEKL and the congenic marker Thy1.1. These cells were adoptively transferred into wild type mice followed by influenza-OVA infection and induction of Atg5 knockout at day 5 after infection. Tracking the kinetic of the cells show, that the Atg5 knockout cells do not expand as much as the control cells and they experience complete contraction within few days. No memory population is established. To investigate differences in Atg5 knockout cells versus control cells we sorted the donor cells at the peak of immune response and investigated their gene expression profile via microarray analysis. Overall design: 10.000 naive CD8 T cells were isolated from the spleens of transgenic mice (untreated) and were adoptively transferred into naive wild type mice. On the next day these mice got intranasally infected with 15 plaque forming units of Influenza strain A/PR8-OVA. At day 5 to 8, Atg5 knockout was induced by intraperitoneal tamoxifen administration. At day 7 and 8, donor cells were sorted from spleens, RNA was isolated and investigated via microarray. Three groups of transgenic cells were compared in two independent experiments: Atg5 knockout cells were compared to two groups of control cells: 1. heterozygously Atg5 loxP expressing cells and 2. homozygously Atg5 loxP expressing cell that lacked the expression of Cre-recombinase.