Figure 4.

Mitochondria-Targeted β-Actin but Not Nucleus-Targeted β-Actin Shows Rescue Effect on MMP

(A) Schematics of retroviral constructs used for re-introducing GFP, β-actin (Actb), β-actin with NLS (ActbNLS), and β-actin with MTS (ActbMTS) into KO (β-actin−/−) MEFs.

(B) qPCR quantification of relative expression level of β-actin mRNA. WT::GFP represents WT cells transduced with retrovirus carrying GFP; n = 3 biological replicates.

(C) Western blot of protein levels of HA-tagged GFP and β-actin. The HA-GFP (first two lanes), HA-tagged β-actin, β-actin with NLS, and β-actin with MTS are indicated by asterisk, with expected molecular weights.

(D) Confocal images of localization of HA-tagged GFP, β-actin, β-actin with NLS, and β-actin with MTS. Scale bar, 8 μm.

(E) FACS analysis of MMP stained by MitoTracker Orange; n = 4 independent experiments. One-way analysis of variance (ANOVA) with Tukey's post hoc test: mean ± SEM, **p ≤ 0.01). SEM, standard error of the mean.

(F) mtDNA level comparison between KO::GFP and KO::ActbMTS cells; n = 3 biological replicates, Student's t test.

(G) Transcript level of MT-ND1, MT-CYB, and MT-CO1 between KO::GFP and KO::ActbMTS cells; n = 3 independent experiments; Student's t test: mean ± SEM *p ≤ 0.05, **p ≤ 0.01.

See also Figure S5.