GTP-locked Irgb10^K81A mutant but not wildtype Irgb10 targets <i>C. trachomatis</i> inclusions in <i>Atg3</i>- and <i>Atg5</i>-deficient cells with high efficiency.

<p>(<b>A</b>) <i>Atg5^−/−</i> MEFs were transfected with GFP-fusion constructs expressing either wildtype Irgb10 (WT) or the Irgb10^S82N mutant that is deficient of GTP binding. Cells were subsequently infected with <i>C. trachomatis</i> and treated with 200 U/ml of IFNγ at 3 hpi. Fixed cells were stained with Hoechst. Representative images are shown. (<b>B</b>) WT, <i>Atg3^−/−</i> & <i>Atg5^−/−</i> MEFs were transfected with the indicated constructs. Cells were infected with <i>C. trachomatis</i> and treated with IFNγ at 3 hpi. Cells were fixed at 20 hpi and stained for the <i>C. trachomatis</i> inclusion membrane marker IncG as well as DNA (Hoechst). Representative images are shown. (<b>C</b>) Graphical representation of the frequency at which WT Irgb10 and the Irgb10^K81A mutant colocalize with inclusions. Average values ± SD of three independent experiments are shown. Differences in the targeting frequency for WT Irgb10 and Irgb10^K81A to inclusions were evaluated for statistical significance (**, p<0.005).