Surface Accessibility and Conformational Changes in the N-terminal Domain of Type I Inositol Trisphosphate Receptors

Anyatonwu, Georgia I.; Joseph, Suresh K.

doi:10.1074/jbc.m806932200

Cited by 8 publications

(3 citation statements)

References 53 publications

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“…7, shown in red). This has been confirmed experimentally (37). It should be noted that the Ca 2ϩ -dependent change in MPEG accessibility of fragment V also was observed when the MPEG reaction was carried out after the receptor had been first cleaved with trypsin (data not shown).…”

Section: Discussionsupporting

confidence: 63%

Calcium-dependent Conformational Changes in Inositol Trisphosphate Receptors

Anyatonwu

Khan

Schug

et al. 2010

Journal of Biological Chemistry

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We have used limited trypsin digestion and reactivity with PEG-maleimides (MPEG) to study Ca 2؉ -induced conformational changes of IP 3 Rs in their native membrane environment. We found that Ca 2؉ decreased the formation of the 95-kDa C-terminal tryptic fragment when detected by an Ab directed at a C-terminal epitope (CT-1) but not with an Ab recognizing a protected intraluminal epitope. This suggests that Ca 2؉ induces a conformational change in the IP 3 R that allows trypsin to cleave the C-terminal epitope. Half-maximal effects of Ca 2؉ were observed at ϳ0.5 M and was sensitive to inhibition by IP 3 . Ca 2؉ also stimulated the reaction of MPEG-5 with an endogenous thiol in the 95-kDa fragment. This effect was eliminated when six closely spaced cysteine residues proximal to the transmembrane domains were mutated (C2000S, C2008S, C2010S, C2043S, C2047S, and C2053S) or when the N-terminal suppressor domain (amino acids 1-225) was deleted. A cysteine substitution mutant introduced at the C-terminal residue (A2749C) was freely accessible to MPEG-5 or MPEG-20 in the absence of Ca 2؉ . However, cysteine substitution mutants in the interior of the tail were poorly reactive with MPEG-5, although reactivity was enhanced by Ca 2؉ . We conclude the following: a) that large conformational changes induced by Ca 2؉ can be detected in IP 3 Rs in situ; b) these changes may be driven by Ca 2؉ binding to the N-terminal suppressor domain and expose a group of closely spaced endogenous thiols in the channel domain; and c) that the C-terminal cytosol-exposed tail of the IP 3 R may be relatively inaccessible to regulatory proteins unless Ca 2؉ is present.Inositol 1,4,5-trisphosphate receptors (IP 3 Rs) 3 are large tetrameric intracellular Ca 2ϩ -release channels that mediate the release of Ca 2ϩ from endoplasmic reticulum stores into the cytosol in response to IP 3 (1-5). Three different isoforms exist, which can form homo-and heteroligomers. All three isoforms consist of an IP 3 binding domain in the N-terminal region, a channel domain containing six transmembrane segments in the C-terminal region and an intervening regulatory domain. Apart from IP 3 , the principal regulator of the channel is Ca 2ϩ , which exerts a biphasic effect on channel function with activation at low concentrations and inhibition at high concentrations. The structural organization of this complex protein and the mechanism by which binding of IP 3 and Ca 2ϩ are linked to channel gating are active areas of investigation.An important feature of the mechanism of the channel involves conformational changes in the protein. In particular, the gating mechanism is thought to involve conformational changes initiated by IP 3 binding that are propagated to various regions of the channel domain (6, 7). The IP 3 binding domain of the receptor is organized into a "core" segment that binds IP 3 with high affinity (amino acids 226 -604) and a "suppressor domain" that inhibits IP 3 binding to the core (amino acids 1-225). Crystal structures for the suppressor domain and the ...

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Section: Discussionsupporting

confidence: 63%

Calcium-dependent Conformational Changes in Inositol Trisphosphate Receptors

Anyatonwu

Khan

Schug

et al. 2010

Journal of Biological Chemistry

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“…1d ), attenuates InsP 3 -evoked Ca 2+ release 8 ; and Ca 2+ , a co-regulator of InsP 3 R 14 , causes the loop to become accessible. 15 The disease-associated ‘hot spot loop’ of RyR1 11 sits at the same location within the ABC structure 9 ( Fig. 1d ) and a mimetic peptide causes RyR2 to become leaky 16 .…”

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confidence: 99%

Structural and functional conservation of key domains in InsP3 and ryanodine receptors

Seo

Velamakanni

Ishiyama

et al. 2012

Nature

166

271

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Inositol 1,4,5-trisphosphate receptors (InsP3R) and ryanodine receptors (RyR) are tetrameric intracellular Ca2+ channels1. For each, the pore is formed by C-terminal transmembrane domains and regulated by signals detected by the large cytosolic structures. InsP3R gating is initiated by InsP3 binding to the InsP3-binding core (IBC, residues 224-604 of InsP3R1)2 and it requires the suppressor domain (SD, residues 1-223)2-8. We present structures of the N-terminal region (NT) of InsP3R1 with (3.6 Å) and without (3.0 Å) InsP3 bound. The arrangement of the three NT domains, the SD, IBC-β and IBC-α, identifies two discrete interfaces (α and β) between the IBC and SD. Similar interfaces occur between equivalent domains (A, B and C) in RyR19. The orientations of the three domains docked into a tetrameric structure of InsP3R10 and of the ABC domains in RyR9 are remarkably similar. The importance of the α-interface for activation of InsP3R and RyR is confirmed by mutagenesis and, for RyR, by disease-causing mutations9,11,12. InsP3 causes partial closure of the clam-like IBC, disrupting the β-interface and pulling the SD towards the IBC. This reorients an exposed SD loop (HS-loop) that is essential for InsP3R activation7. The loop is conserved in RyR and includes mutations associated with malignant hyperthermia and central core disease9,11,12. The HS-loop interacts with an adjacent NT, suggesting that activation re-arranges inter-subunit interactions. The A-domain of RyR functionally replaced the SD in a full-length InsP3R, and an InsP3R in which its C-terminal transmembrane region was replaced by that from RyR1 was gated by InsP3 and blocked by ryanodine. Activation mechanisms are conserved between InsP3R and RyR. Allosteric modulation of two similar domain interfaces within an N-terminal subunit re-orients the first domain (SD or A-domain), allowing it, via interactions of the second domain of an adjacent subunit (IBC-β or B-domain), to gate the pore.

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“…Two intrinsic determinants stabilizing the subunit configurations can be considered a priori-IP 3 and Ca 2+ -because channel activation requires both IP 3 and Ca 2+ . In fact, the recent X-ray structures of the IBD indicate that conformational changes occur upon IP 3 binding (7,8); our group and others have demonstrated Ca 2+ -induced structural changes (84,85,94,95). Fluorescence resonance energy transfer (FRET) analysis of recombinant IP 3 Rs also showed reversible structural changes induced by physiological concentrations of IP 3 and Ca 2+ (96).…”

Section: Discussionmentioning

confidence: 90%

Aberrant calcium signaling by transglutaminase-mediated posttranslational modification of inositol 1,4,5-trisphosphate receptors

Hamada

Terauchi

Nakamura

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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The inositol 1,4,5-trisphosphate receptor (IP 3 R) in the endoplasmic reticulum mediates calcium signaling that impinges on intracellular processes. IP 3 Rs are allosteric proteins comprising four subunits that form an ion channel activated by binding of IP 3 at a distance. Defective allostery in IP 3 R is considered crucial to cellular dysfunction, but the specific mechanism remains unknown. Here we demonstrate that a pleiotropic enzyme transglutaminase type 2 targets the allosteric coupling domain of IP 3 R type 1 (IP 3 R1) and negatively regulates IP 3 R1-mediated calcium signaling and autophagy by locking the subunit configurations. The control point of this regulation is the covalent posttranslational modification of the Gln2746 residue that transglutaminase type 2 tethers to the adjacent subunit. Modification of Gln2746 and IP 3 R1 function was observed in Huntington disease models, suggesting a pathological role of this modification in the neurodegenerative disease. Our study reveals that cellular signaling is regulated by a new mode of posttranslational modification that chronically and enzymatically blocks allosteric changes in the ligand-gated channels that relate to disease states.L igand-gated ion channels function by allostery that is the regulation at a distance; the allosteric coupling of ligand binding with channel gating requires reversible changes in subunit configurations and conformations (1). Inositol 1,4,5-trisphosphate receptors (IP 3 Rs) are ligand-gated ion channels that release calcium ions (Ca 2+ ) from the endoplasmic reticulum (ER) (2, 3). IP 3 Rs are allosteric proteins comprising four subunits that assemble a calcium channel with fourfold symmetry about an axis perpendicular to the ER membrane. The subunits of three IP 3 R isoforms (IP 3 R1, IP 3 R2, and IP 3 R3) are structurally divided into three domains: the IP 3 -binding domain (IBD), the regulatory domain, and the channel domain (3-6). Fitting of the IBD X-ray structures (7, 8) to a cryo-EM map (9) indicates that the IBD activates a remote Ca 2+ channel by allostery (8); however, the current X-ray structure only spans 5% of each tetramer, such that the mechanism underlying allosteric coupling of the IBD to channel gating remains unknown.The IP 3 R in the ER mediates intracellular calcium signaling that impinges on homeostatic control in various subsequent intracellular processes. Deletion of the genes encoding the type 1 IP 3 R (IP 3 R1) leads to perturbations in long-term potentiation/ depression (3, 10, 11) and spinogenesis (12), and the human genetic disease spinocerebellar ataxia 15 is caused by haploinsufficiency of the IP 3 R1 gene (13-15). Dysregulation of IP 3 R1 is also implicated in neurodegenerative diseases including Huntington disease (HD) (16-18) and Alzheimer's disease (AD) (19)(20)(21)(22). IP 3 Rs also control fundamental cellular processes-for example, mitochondrial energy production (23, 24), autophagy regulation (24-27), ER stress (28), hepatic gluconeogenesis (29), pancreatic exocytosis (30), and macrophag...

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Surface Accessibility and Conformational Changes in the N-terminal Domain of Type I Inositol Trisphosphate Receptors

Cited by 8 publications

References 53 publications

Calcium-dependent Conformational Changes in Inositol Trisphosphate Receptors

Calcium-dependent Conformational Changes in Inositol Trisphosphate Receptors

Structural and functional conservation of key domains in InsP3 and ryanodine receptors

Aberrant calcium signaling by transglutaminase-mediated posttranslational modification of inositol 1,4,5-trisphosphate receptors

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