Nutrient sensing and cell fate determination through structural allostery in PASK - Project Summary Nutrient sensing is fundamental to linking environmental conditions with cellular energy balance and fate decisions—processes that are central to human health and, when deregulated, contribute to diseases such as cancer, diabetes, tissue degeneration, and aging. Nutrient-sensitive kinases such as mTOR, GCN2, AMPK, and PASK play pivotal roles in maintaining cellular homeostasis by dynamically responding to environmental cues. Among these, the Per-Arnt-Sim (PAS) domain kinase (PASK) is particularly notable due to its unique monomeric architecture. It combines metabolite-sensing PAS domains with a serine/threonine kinase domain, enabling it to directly interpret metabolic and nutrient signals. This streamlined design allows PASK to translate environmental inputs into precise cellular responses that regulate metabolism, growth, and differentiation. Despite its critical role, the mechanisms by which PAS domains detect environmental cues and transmit this information to the kinase domain remain poorly understood. Through structural phylogenetic analysis, we recently revised the domain architecture of PASK. In addition to the previously known PAS-A domain, we identified two additional PAS domains—PAS-B and PAS-C—within its primary sequence. Notably, the PAS-C domain comprises two subdomains: the PAS-fold and the PAC-motif, separated by a long, unstructured regulatory loop—an architecture that is conserved across metazoans. This loop serves as a target for nutrient signals, driving the dynamic assembly of the PAS-C domain. This assembly, in turn, organizes PASK’s quaternary structure and activates its catalytic function, revealing a novel mechanism of environmental sensing through structural allostery. Building on this discovery, we aim to explore how the PAS domains coordinate with the kinase domain to regulate PASK’s activity, stability, and substrate specificity. In Aim 1, we will map inter- and intra-domain amino acid contact sites across the PASK quaternary structure and examine how these are remodeled in response to signaling cues and are exploited by disease-causing variants to lock PASK in a constitutively active or inactive state. In Aim 2, we will investigate a metabolically regulated acetylation-ubiquitylation switch that targets the PAS domain and controls PASK stability during the stem cell cycle, influencing the balance between self-renewal and differentiation. In Aim 3, we will characterize how nutrient signals flow through the PAS domains to phosphorylate newly identified PASK-interacting partners and substrates in mammalian stem cells. By integrating structural biology, biochemistry, and stem cell biology, this project aims to define the molecular mechanisms by which PASK regulates cell behavior and metabolic processes. These insights will illuminate the pathogenesis of metabolic syndrome and tissue dysfunction observed in PASK-deficient models, potentially opening new avenues for therapeutic intervention.
