Skeletal muscle homeostasis depends on the precise coordination of autophagy and mitochondrial quality control, yet the long noncoding RNA (lncRNA) networks governing these processes remain poorly characterized. Here, we identify Wee1-AS as a differentiation-induced lncRNA whose expression is markedly upregulated during C2C12 myogenesis. Antisense oligonucleotide (ASO)-mediated depletion of Wee1-AS caused pronounced accumulation of the autophagy receptor p62/SQSTM1 and LC3-II, accompanied by a reduction in MyHC2a, indicative of impaired myotube maturation. Mechanistically, loss of Wee1-AS severely disrupted lysosomal acidification and blocked the proteolytic maturation of Cathepsin D and Cathepsin L, thereby preventing productive autophagosome–lysosome fusion and arresting autophagic flux. This lysosomal insufficiency propagated a cascade of mitochondrial dysfunction, manifested by dissipated membrane potential, diminished succinate dehydrogenase (SDH) activity, and broad downregulation of oxidative phosphorylation (OXPHOS) complexes. Collectively, our findings establish Wee1-AS as an essential upstream regulator of lysosomal degradative capacity in skeletal muscle, whose activity is indispensable for coupling autophagic flux to mitochondrial health. These results suggest Wee1-AS as a potential therapeutic target in metabolic myopathy conditions associated with autophagic and mitochondrial failure.