Fig. 2. MYTHO is required for skeletal muscle autophagy.

A Representative confocal microscope images were used for the quantification of the colocalization of MYTHO-GFP and cherry-LC3B (upper panels) or MYTHO-GFP and LAMP2-cherry (lower panels) in isolated FDB muscle fibers. Yellow dots highlight colocalization. Scale bar: 20 µm. B Quantification of MYTHO positive puncta that co-localizes with LC3 in single-fibers from mice treated with colchicine or vehicle in fed (FED) and 24 h starved (STV). C Quantification of MYTHO-positive puncta co-localizing with LAMP2-cherry in fed (FED) and 24 h starved (STV) mice that were treated with colchicine or vehicle. D Representative images of single FDB fibers from FED or 24 h STV mice treated with colchicine or vehicle that were co-transfected with sh-RNAs against Mytho or scrambled (scrmb) together with cherry-LC3B. Scale bar: 20 µm. E LC3 positive puncta/area were quantified in FED and 24 h STV single FDB fibers from mice treated with colchicine or vehicle for flux measurements (n = 3 mice per condition). Data in B, C, and E were analyzed with two-way ANOVA, and corrections for multiple comparisons were performed with the two-stage step-up method of Benjamini, Krieger, and Yekutieli (∗p < 0.05 and q < 0.1). Data are presented as mean ± SEM (with individual data points). Detailed information on raw data, statistical tests, p values, and q values are provided in the Source Data file. The drawings in A and B were created with BioRender.com.