ADORA2B/ador-1 is required for multi-system transcriptional adaptation to mild hypoxia and healthspan extension in C. elegans

BACKGROUND: Mild intermittent hypoxia is emerging as a promising strategy to enhance healthspan, but the molecular mechanisms remain poorly defined. ADORA2B, a hypoxia-inducible adenosine receptor, is known to regulate metabolism and stress responses, yet its role in functional aging is unclear. METHODS: Using Caenorhabditis elegans, we investigated the role of the ADORA2B ortholog ador-1 in mediating the healthspan effects of mild hypoxia. We exposed wild-type and ador-1 knockout worms to low-dose cobalt chloride (CoCl₂) and assessed movement, speed, and lifespan metrics through automated imaging. Transcriptomic analyses via 3' RNA-seq and Gene Set Enrichment Analysis (GSEA) were performed to characterize underlying molecular responses. RESULTS: Mild hypoxia significantly extended healthspan in wild-type C. elegans, increasing sustained locomotor activity and distance traveled throughout adulthood. This benefit was markedly attenuated in ador-1 mutants, which displayed reduced baseline healthspan and a substantially blunted transcriptional response to CoCl₂ treatment. Transcriptomic analyses identified coordinated regulation of canonical hypoxia-responsive genes, including acs-2, icl-1, adh-1, ftn-1 and ftn-2, consistent with activation of hypoxia-responsive transcriptional programs. GSEA revealed activation of pathways related to neuronal plasticity, muscle function and mitochondrial adaptation, together with transcriptional downregulation of ROS-related pathways. In contrast, hypoxia-treated ador-1 mutants exhibited suppression of pathways associated with ciliary function, immune responses, protein synthesis and cellular homeostasis. CONCLUSIONS: ADORA2B is required for the coordinated transcriptional adaptation associated with hypoxia-induced healthspan extension in C. elegans. Loss of ador-1 is associated with impaired baseline healthspan and an attenuated response to hypoxic conditioning. These findings support a model in which successful hypoxic adaptation requires coordinated regulation of multiple biological systems.