Alternative mitophagy is defective in ulk1cKO mice during TAC. (A–E) 3–4-month-old WT and ulk1cKO mice were subjected to either sham operation or TAC for 3 days. Heart lysates were analysed by Western blotting using anti-Rip1, anti-Rip3, anti-phospho-Rab9 (S179), anti-Rab9, anti-Ulk1, and anti-αTubulin antibodies (A). The relative ratios of phospho-Rab9 to Rab9 (B) were quantified. Mean values ± S.E., **P < 0.01. Statistical analyses were conducted with two-way ANOVA followed by Tukey’s post hoc test. (C–E) Samples from hearts of mice subjected to 3-day TAC were subjected to electron microscopic analyses. Representative EM images are shown in upper panel of (C). Enlarged mitochondria are shown in lower panel of (C). Mitochondria aspect ratio was quantified. Mean values ±S.E., *P < 0.05. Statistical analyses were conducted with two-way ANOVA followed by Šídák’s multiple comparisons test (D). Mitochondria were categorized by the morphology of cristae and quantified. The percentage of structured or irregular cristae and empty mitochondria was quantified (n = 5 each group) (E).