RESUMEN
While young muscle is capable of restoring the original architecture of damaged myofibers, aged muscle displays a markedly reduced regeneration. We show that expression of the "anti-aging" protein, α-Klotho, is up-regulated within young injured muscle as a result of transient Klotho promoter demethylation. However, epigenetic control of the Klotho promoter is lost with aging. Genetic inhibition of α-Klotho in vivo disrupted muscle progenitor cell (MPC) lineage progression and impaired myofiber regeneration, revealing a critical role for α-Klotho in the regenerative cascade. Genetic silencing of Klotho in young MPCs drove mitochondrial DNA (mtDNA) damage and decreased cellular bioenergetics. Conversely, supplementation with α-Klotho restored mtDNA integrity and bioenergetics of aged MPCs to youthful levels in vitro and enhanced functional regeneration of aged muscle in vivo in a temporally-dependent manner. These studies identify a role for α-Klotho in the regulation of MPC mitochondrial function and implicate α-Klotho declines as a driver of impaired muscle regeneration with age.
Asunto(s)
Envejecimiento/genética , ADN Mitocondrial/genética , Mitocondrias/genética , Músculo Esquelético/metabolismo , Mioblastos/metabolismo , Receptores de Superficie Celular/genética , Células Madre/metabolismo , Envejecimiento/metabolismo , Envejecimiento/patología , Animales , Metilación de ADN , ADN Mitocondrial/metabolismo , Epigénesis Genética , Regulación del Desarrollo de la Expresión Génica , Glucuronidasa , Proteínas Klotho , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias/metabolismo , Músculo Esquelético/patología , Mioblastos/patología , Regiones Promotoras Genéticas , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Receptores de Superficie Celular/antagonistas & inhibidores , Receptores de Superficie Celular/metabolismo , Regeneración/genética , Transducción de Señal , Células Madre/patologíaRESUMEN
Mitophagy is critical for cell homeostasis. Externalization of the inner mitochondrial membrane phospholipid, cardiolipin (CL), to the surface of the outer mitochondrial membrane (OMM) was identified as a mitophageal signal recognized by the microtubule-associated protein 1 light chain 3. However, the CL-translocating machinery remains unknown. Here we demonstrate that a hexameric intermembrane space protein, NDPK-D (or NM23-H4), binds CL and facilitates its redistribution to the OMM. We found that mitophagy induced by a protonophoric uncoupler, carbonyl cyanide m-chlorophenylhydrazone (CCCP), caused externalization of CL to the surface of mitochondria in murine lung epithelial MLE-12 cells and human cervical adenocarcinoma HeLa cells. RNAi knockdown of endogenous NDPK-D decreased CCCP-induced CL externalization and mitochondrial degradation. A R90D NDPK-D mutant that does not bind CL was inactive in promoting mitophagy. Similarly, rotenone and 6-hydroxydopamine triggered mitophagy in SH-SY5Y cells was also suppressed by knocking down of NDPK-D. In situ proximity ligation assay (PLA) showed that mitophagy-inducing CL-transfer activity of NDPK-D is closely associated with the dynamin-like GTPase OPA1, implicating fission-fusion dynamics in mitophagy regulation.