rragc KI fish manifests reduced TFEB activity and defective function of lysosomal clearance. (A) Heatmap of relative mRNA expression levels of genes regulated by TFEB in rragc KI and WT fish hearts. Values are shown as fold change of relative mRNA expression normalized to the mean value of WT. actb2 was used as an internal control. lamp1, lysosomal associated membrane protein 1; atg9a, autophagy related 9A; atg9b, autophagy related 9B; uvrag, UV radiation resistance associated gene; vps11, VPS11 core subunit of CORVET and HOPS complexes. (B) Immunoblot of LC3 in heart lysates of 9-month-old rragc KI and WT fish treated with BafA1 (1.2 ug/g, 6 h) or vehicle via intraperitoneal injection. β-actin as a loading control. * p < 0.05 versus WT. (C) Total lysate and insoluble protein fractions of rragc KI and WT fish heart were separated by SDS–PAGE and immunoblotted against ubiquitin antibodies (Ub). Fish age around 1 year. L.E., longer exposure. (D) Periodic Acid–Schiff (PAS) staining of fish heart tissues. Representative images are shown. Scale bars, 30 μm. The right panel is the higher magnification views of sections indicated by white dash box. Arrows point to glycogen deposit.
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