Cryo-EM insights into isoform-specific properties of the IP₃R2 channel

Calcium release through inositol 1,4,5-trisphosphate receptors (IP₃Rs) is a fundamental signaling mechanism that regulates diverse cellular processes. Among the three mammalian IP₃R isoforms, IP₃R2 is widely expressed, yet its structural basis for activation and regulation remains unclear. Here, we report cryo-EM structures of mammalian IP₃R2 in ligand-free (closed) and Ca^2+/IP₃/ATP-bound (activated) states at 3.3 Å and 3.6 Å resolution, respectively. These structures define the architecture of IP₃R2 and reveal conformational transitions associated with channel activation. Although the IP₃-binding pocket is conserved, subtype-specific differences in IP₃ affinity likely arise from conformational dynamics of the regulatory ARM2 domain. Comparative analyses of IP₃R isoforms identify subtype-specific allosteric networks and domain motions that underlie differential regulation. We further define the ATP-binding site and, through mutagenesis and electrophysiology, establish the structural basis for ATP modulation of channel activity. Together, these findings reveal mechanisms of IP₃R2 activation and subtype-specific regulation, providing a framework for understanding isoform-dependent Ca²⁺ signaling.