PROJECT SUMMARY / ABSTRACT A fundamental challenge in our rapidly aging society is how to extend healthspan, or maintain robust fitness without major diseases. The circadian clock, our intrinsic timer, is a pivotal cellular mechanism to promote health and healthy aging, as both loss- and gain-of-function studies have provided strong experimental evidence for a causative role of circadian rhythms in aging. This is important because in humans, environmental circadian disruptions associated with modern 24/7 lifestyles also aggravate disease risk and age-related decline. Despite growing functional insights, a significant gap in knowledge remains regarding cellular mechanisms and interventional strategies that impinge on circadian timing to promote healthy aging. The ROR (Retinoid acid receptor-related Orphan Receptor) subfamily of nuclear receptors are key clock components, functioning to sustain oscillatory robustness and phase which is known to alter with age. We previously identified an agonist of RORs called Nobiletin (NOB) and demonstrated a strong efficacy of NOB to improve physiological fitness in disease models and aged mice. Extending the pharmacological gain-of-function studies, we recently generated muscle-specific double knockout of Rora and Rorc in mice (Ror mDKO), and found that adult and aged Ror mDKO mice showed impaired skeletal muscle function and mitochondrial dysfunction in a circadian time- dependent manner. Importantly, circadian transcriptomic analysis showed significantly altered circadian expression (level, amplitude and phase) of OXPHOS genes. We hypothesize that RORs control mitochondrial respiration and skeletal muscle function via circadian transcriptional and epigenetic mechanisms, and play a modifiable role in lifestyle interventions to enhance healthy aging. In Aim 1, we will comprehensively investigate the role of RORs in age- and circadian time-dependent changes in mitochondrial activity and homeostasis in skeletal muscle. In Aim 2, we will determine circadian transcriptional and epigenetic mechanisms by RORs in skeletal muscle, especially in the concerted regulation of OXPHOS gene expression. In Aim 3, combining timed exercise and NOB treatment, we will determine the requisite role of RORs in circadian interventions to prolong healthspan and lifespan. Together, the proposed studies will provide key mechanistic insights into circadian regulation of skeletal muscle mitochondria, and pinpoint RORs as an actionable target for lifestyle interventions to enhance healthy aging. The innovations include new insights into the function and mechanisms of RORs in skeletal muscle aging, a novel tissue-specific double KO mouse line and circadian epigenetic/3D genome studies, and versatile circadian interventions. We have assembled an excellent investigator team with complementary expertise and demonstrated track records in aging, circadian rhythms, mitochondrial metabolism, genetic/epigenetic regulation, and interventions. Given ...