Fig. 6

Neuronal Nr4a1 defect is an endogenous signal critical to the synaptic location of C1q in MRL/lpr mice. a Heat maps showing the relative expression of significantly altered genes (involved in Nr4a signaling) generated from the hippocampal RNA sequencing of MRL/mpj vs. MRL/lpr mice at 6 weeks of age. Representative PCR and quantification of Nr4a1 mRNA levels in brain lysates from 6-week-old MRL/mpj and MRL/lpr (n = 5) mice (b) and hippocampal lysates from both 6- and 16-week-old mice (c). The relative expression was normalized to the average of MRL/mpj controls. d Representative images of dendritic segments and quantification of C1q colocation after treatment of hippocampal neurons with the indicated shRNA or shRNA-plus-phallacidin treatment for 3 days. Scale bar, 20 μm. At least 15 neurons per culture from three independent cultures were used for the analysis. e and f Dendritic segments of neurons after treatment with control or Nr4a1-specific shRNA and incubation with phallacidin or the vehicle (e). Scale bars as indicated. The histogram shows the spine density along dendrites after treatment with shRNAs plus or minus phallacidin, as indicated (f). At least 11 neurons per culture from three independent neuronal cultures were used for the analysis. g and h Representative images of F-actin and PSD-95 in the hippocampal CA3 region. (h) Quantitation of the intensity of the F-actin signal overlapping PSD-95 puncta. Scale bar, 10 μm. Each dot represents the average for one mouse, with 7–8 mice per group. Data shown are the mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; and ns, not significant by one-way ANOVA with Tukey’s post hoc test in (b), (f) and unpaired Student’s t-test in (c), (d), (h). See also Supplementary Fig. 8