FIGURE 5.
Top, AlphaScreenTM analysis of the interaction between TLR4ic mutants and TRAM. HEK293T cells were transiently co-transfected with plasmids encoding the E-tagged TLR4 WT or mutants and FLAG-tagged TRAM. The transfected cells were lysed after 48 h, and protein interactions were detected with the AlphaScreen FLAGTM (M2) detection kit (PerkinElmer Life Sciences). The results are displayed as fold induction, namely relative to the counts of an irrelevant E-tagged construct. Bottom, expression levels of the E-tagged TLR4 constructs were detected on Western blot; TLR4 is the lower band on the blot. In the 1st lane (negative control (neg. ctrl)), the control lysate was loaded; the order of the other loaded samples aligns with the order of the samples on the graph.