Figure 3.

Alterations of mitochondrial functions in plectin isoform-specific knockout tissue. (A and B) Cross-sections of soleus muscle obtained from 12-week-old isoform-specific P1b (A) and P1d (B) knockout mice were stained for SDH. Boxed areas are shown enlarged. Scale bars: 50 µm (left panels) and 25 µm (right panels). (C) Equal amounts of wild-type, P1b-KO and P1d-KO gastrocnemius muscle lysates were subjected to immunoblotting using antibodies as indicated. GAPDH, loading control. (D and E) Signal intensities of immunoblots as shown in (B) were densitometrically measured and normalized to total protein content as analyzed by the Coomassie staining (not shown). Mean values ± SEM, three experiments. Note that the reduced protein levels observed in P1b-KO and P1d-KO lysates were statistically significant [Complex II, P < 0.01 (P1b-KO) and P < 0.001 (P1d-KO); Complex IV, P < 0.05 (P1b-KO) and P < 0.01 (P1d-KO)]. (F) CS activity was measured in wild-type, P1b-KO and P1d-KO gastrocnemius muscle lysates prepared from 12-week-old mice. Mean values ± SEM, four experiments. (G and H) Relative protein levels as assessed in (C) were normalized to respective CS activity levels as determined in (F). Note that the overall protein levels of respiratory complex subunit proteins per mitochondrion remained unchanged in P1b-KO and P1d-KO muscles. (I) Respiratory capacities of mitochondria in permeabilized muscle fibers isolated from heart, soleus or gastrocnemius muscles of wild-type, P1b-KO and P1d-KO mice. Mean ± SD, three experiments. (J) Apparent K_m for ADP in permeabilized muscle fibers isolated from heart, soleus and gastrocnemius muscles of mouse lines as indicated. Mean ± SD, three experiments.