Fig. 6

LY6E regulated the IFN-α- and IC-induced release of mtDNA into the cytosol. BMDMs were electroporated with siLY6E or siCtl and then stimulated with or without IFN-α or ICs for 24 h. Both total DNA and cytosolic DNA were extracted to measure mtDNA levels with specific primers via qPCR, as described in the Materials and Methods. The relative abundance of total and cytosolic mtDNA was determined by normalization to actin or an exogenously added plasmid encoding the FLAG gene (PCR3.1-flag) (A). The levels of cytosolic mtRNA were similarly measured (B). The expression of two mtRNA downstream signaling molecules, RIG-1 and MDA5, was determined by qPCR (C). Several mtDNA downstream signaling molecules, such as STING and TBK1, were examined for their activation status in IFN-α-stimulated (D) and IC-stimulated (E) BMDMs by Western blotting. For Western blotting, the samples were derived from the same experiment, and both the gels and the blots were processed in parallel. Each data point represents one mouse, and the values are fold changes relative to the mean value of the siCtl in RT‒qPCR and Western blotting. Two-way ANOVA with Holm‒Sidak multiple comparisons was used to compare differences among different treatments. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001