Fig. 2: Chka protein expression is inversely correlated with the rostro-caudal gradient of severity in Chkb-mediated muscular dystrophy.

Transmission electron microscopy (TEM) appearance of A forelimb (triceps) and B hindlimb (quadriceps) of 115-day old Chkb^−/− mice showing extensive injury in hindlimb not the forelimb. A, B are representative of 3 mice per group with similar appearance. Scale bar = 1 μm. Western blot of C forelimb (triceps) and D hindlimb (quadriceps) samples from three distinct (lanes 1–3) Chkb^+/+, four distinct (lanes 4–7) Chkb^+/− and three distinct (lanes 8–10) Chkb^−/− mice probed with anti-Chka, anti-Chkb, and anti-Gapdh antibodies. Bottom: densitometry of the WB data shows the ratio of Chka and Chkb to Gapdh. Chka signal is not significantly different in forelimb and hindlimb samples from Chkb^+/− mice compared to the wild type. Chka is upregulated in forelimb muscles and downregulated in hindlimb muscles from Chkb^−/− mice. Chkb signal is decreased in hindlimb and forelimb muscle samples of Chkb^+/− mice and is absent in muscle samples of Chkb^−/− mice. Values in C and D are means ± SD; For C and D, n = 3 independent Chkb^+/+, 4 independent Chkb^+/− and 3 independent Chkb^−/− mice per group. For C, left, one-way ANOVA with Tukey’s multiple comparison test, p = 0.04335. For C, right, two-sided student’s t-test, p = 0.02842. For D, left, one-way ANOVA with Tukey’s multiple comparison test, p = 0.00011. For D, right, two-sided student’s t-test, p = 0.01553. E Transmission electron microscopy (TEM) appearance of the mitochondrial profile of hindlimbs from 12-day old and 60 days old wild-type (Chkb^+/+) and Chkb-deficient (Chkb^−/−) mice. For E, images are representative of 3 mice per group with similar appearance. Scale bar = 300 nm. F At 12 days of age hindlimbs from wild type and Chkb^−/− mice had the same number of mitochondria per imaged field however, the volume density of the Chkb^−/− mitochondria was increased and the cristae density was preserved. At 115 days of age, Chkb^−/− mitochondria were fewer in number, had markedly reduced cristae density, and were much larger in size. The increased size of the mitochondria at this age accounts for the preserved volume density. Values in F are means ± SEM. For F, top, n = 9 (12 days old Chkb^+/+), n = 6 (12 days old Chkb^−/−), n = 10 (60 days old Chkb^+/+) and n = 6 (60 days old Chkb^−/−) muscle sections from 3 independent mice per genotype for each timepoint. For F, middle, n = 7 (12 days old Chkb^+/+), n = 6 (12 days old Chkb^−/−), n = 5 (60 days old Chkb^+/+), n = 7 (60 days old Chkb^−/−) muscle sections from 3 independent mice per genotype for each timepoint. For F, bottom, n = 7 (12 days old Chkb^+/+), n = 6 (12 days old Chkb^−/−), n = 8 (60 days old Chkb^+/+) and n = 8 (60 days old Chkb^−/−) muscle sections from 3 independent mice per genotype for each timepoint. Two-sided student’s t-test. For (F, top), p = 0.39308, p = 0.0000. For F, middle, p = 0.0077, p = 0.427594. For F, bottom, p = 0.78406, p = 0.00273. * and # p < 0.05, **p < 0.01. Source data are provided as a Source Data file.