Adenosine 5'-monophosphate-activated protein kinase (AMPK), a highly conserved energy-sensing kinase, plays a central role in maintaining cellular metabolic homeostasis, such as glucose/lipid metabolism, mitochondrial biogenesis, and autophagy, to counteract energetic stress. Beyond the canonical adenylate-dependent activation of AMPK by sensing the ATP:AMP/ADP ratio, emerging evidence reveals noncanonical activation mechanisms. Proper AMPK signaling is essential for physiological functions, whereas its dysregulation underlies various pathologies, including metabolic diseases, immune disorders, and cancer. This review systematically recapitulates the historical background and landmark discoveries of the AMPK field, including the discovery of AMPK, the structure of AMPK complex, upstream regulators and downstream substrates, and the transcriptional/posttranscriptional and posttranslational regulatory mechanisms. Furthermore, we delineate its roles in maintaining metabolic equilibrium, mitochondrial integrity, and immune competence across organ systems and highlight how dysregulated AMPK activity contributes to pathological diseases, encompassing alterations in its expression levels, intrinsic kinase activity, subcellular localization or upstream regulatory signaling cascades. Specifically, we focus on knock-in or knockout mouse models utilized to study AMPK-associated human diseases and the translational potential of AMPK-targeting agonists or antagonists across diverse pathological conditions. A deep understanding of the multidimensional regulatory network of AMPK and mechanistic insights with therapeutic innovations that we provide with this review will help address the challenges of complex metabolic diseases.
