Structure of the Type 1 Inositol 1,4,5-Trisphosphate Receptor Revealed by Electron Cryomicroscopy

Abstract

The three-dimensional structure of the type 1 inositol 1,4,5-trisphosphate receptor (InsP 3 R1) has been determined by electron cryomicroscopy and single-particle reconstruction. The receptor was immunoaffinity-purified and formed functional InsP 3 -and heparin-sensitive channels with a unitary conductance similar to native InsP 3 Rs. The channel structure exhibits the expected 4-fold symmetry and comprises two morphologically distinct regions: a large pinwheel and a smaller square. The pinwheel region has four radial curved spokes interconnected by a central core. The InsP 3 -binding core domain has been localized within each spoke of the pinwheel region by fitting its x-ray structure into our reconstruction. A structural mapping of the amino acid sequences to several functional domains is deduced within the structure of the InsP 3 R1 tetramer.The inositol 1,4,5-trisphosphate receptor (InsP 3 R) 1 is a ligand-gated ion channel that modulates Ca 2ϩ release from intracellular stores. The InsP 3 R is widely distributed in metazoans and plays a key role in diverse physiological functions. In mammals, three different InsP 3 R isoforms are expressed; each is encoded by a distinct gene. The three isoforms are homologous and share ϳ70% amino acid identity. Functional channels are composed of four subunits that tetramerize via determinants localized within the transmembrane regions and the cytosolic C terminus (1-3). Individual cell types often express more than one isoform, which may be present as homo-or heterotetrameric populations. The most characterized type 1 InsP 3 R (InsP 3 R1) forms largely homotetramers in cerebellum with M r of 313,000 (2749 residues) for the monomer. Based on biochemical, molecular biological, and electrophysiological studies, each monomer subunit of the InsP 3 R1 is organized into three principal functional regions: an N-terminal InsP 3 -binding region (residues 226 -578), a central (ϳ1,700-residue) modulatory/coupling region, and a channel-forming region near the C terminus. The channel region contains six transmembrane segments interspersed within residues 2276 -2590 that are critical for membrane targeting, oligomerization, and formation of the ion permeation pathway (1-4). Thus, both ends, the large N-terminal region (residues 1-2275) and the C-terminal region (residues 2590 -2749), are exposed to the cytoplasm.Electron micrographs of the negatively stained detergentsolubilized InsP 3 R1 and freeze-fracture deep etching electron microscopy of endoplasmic reticulum have been used to characterize the morphology of the InsP 3 R1 channel (5, 6). But the three-dimensional shape and dimensions of the channel particles were not determined. Two three-dimensional maps of the InsP 3 R1 were reported recently based on single-particle reconstruction from specimens prepared by ice embedding (7) and negative staining (8). There is a poor agreement between these two maps with respect to the overall appearance and dimensions of the channel structure. In this study, we present a 30-Å three-dimensional s...

Three-dimensional Rearrangements within Inositol 1,4,5-Trisphosphate Receptor by Calcium

“…Recent studies have also demonstrated the square-like (29) and the windmill-like three-dimensional maps (28,30). These two structures strongly support our hypothesis of global conformational changes within IP 3 R. The comparison between the S-state and W-state suggests that the structural transition from square to windmill is caused by the relocation of functional domains.…”

“…Accumulating reports have explicitly demonstrated two distinct structures of IP 3 R. One is square-shaped (26,27,29), and the other is windmill-shaped (24,27,30). We have postulated that these dual structures are attributable to global conformational changes between the square state (S-state) and windmill state (W-state) within tetrameric IP 3 R particles as described previously (27).…”

“…From a structural perspective, an investigation of this coupling problem requires information regarding the three-dimensional arrangement of these functional modules. However, knowledge of the overall quaternary structure has been limited because of the apparent heterogeneity within IP 3 R particles (24 -30); thus, the arrangement of functional domains remains controversial.Accumulating reports have explicitly demonstrated two distinct structures of IP 3 R. One is square-shaped (26,27,29), and the other is windmill-shaped (24,27,30). We have postulated that these dual structures are attributable to global conformational changes between the square state (S-state) and windmill state (W-state) within tetrameric IP 3 R particles as described previously (27).…”

IP3 Receptors: Toward Understanding Their Activation

“…Functional IP 3 Rs are tetrameric, assembled either from identical subunits or from mixtures of the three subtypes and their many splice variants Foskett et al 2007). Several structures of the entire IP 3 R1 have been published, each derived from single particle analysis of images from electron microscopy (Hamada and Mikoshiba 2002;Jiang et al 2002a;da Fonseca et al 2003;Hamada et al 2003;Serysheva et al 2003;Sato et al 2004). These studies confirm the tetrameric state of IP 3 R, but variability between the structures and their relatively low resolution ( 30 Å ) have, so far, limited any realistic interpretation of the structural basis of IP 3 R activation (Fig.…”

Structure of TRPV1 channel revealed by electron cryomicroscopy

“…Other eukaryotic homotetrameric ion channels have a twolayer architecture with distinct cytoplasmic and membraneembedded domains, including potassium channels, the InsP 3 receptor, and the ryanodine receptor (28)(29)(30)(31)(32)(33).…”