Three-Dimensional Localization of the α and β Subunits and of the II-III Loop in the Skeletal Muscle L-type Ca2+ Channel

Szpyt, John; Lorenzon, Nancy M.; Pérez, Claudio F.; Norris, Ethan; Allen, Paul D.; Beam, Kurt G.; Samsó, Montserrat

doi:10.1074/jbc.m112.419283

Cited by 12 publications

(25 citation statements)

References 71 publications

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“…2F), engineered at 1a's N terminus. 25 This localization is compatible with previous antibody labeling. 29,30 The atomic structure of the core region of the 2a isoform, encompassing the Src Homology 3 (SH3) domain and a guanylate kinase (GK) domain, 35 with 84% identity with 1a, was docked into the 3D reconstruction of the DHPR complex obtained by TEM taking several factors into account.…”

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confidence: 74%

“…The 3D structures of skeletal DHPR that have remained consistent with the increase in resolution 25,29,30 show a main globular body with an appendage emerging from it. The structural description will be centered in our negative staining 3D reconstruction, which has the highest resolution to date (19 Å) and enabled placing of the 1s and 1a subunits for the first time.…”

mentioning

confidence: 66%

“…The structural description will be centered in our negative staining 3D reconstruction, which has the highest resolution to date (19 Å) and enabled placing of the 1s and 1a subunits for the first time. 25 The DHPR used to obtain these images had a heterologous 1a subunit that contained an YFP to aid in 3D sub-localization and a biotin-acceptor domain tag that was used for purification; the engineered 1a subunit was expressed in transgenic mouse as indicated above. After purification from the muscle tissue, the DHPR sample was negatively stained, imaged by TEM (Fig.…”

mentioning

confidence: 99%

“…The SH3 in the N' region of the subunit was placed adjacent to the identifiable YFP engineered in the N' region. Finally, the two prominent lobes of the GK domain were fit with the two outward spikes in the 3D volume of DHPR 25 ( Fig. 2E-F).…”

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confidence: 99%

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3D structure of muscle dihydropyridine receptor

Samsó¹

2015

Eur J Transl Myol

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Excitation contraction coupling, the rapid and massive Ca 2+ release under control of an action potential that triggers muscle contraction, takes places at specialized regions of the cell called triad junctions. There, a highly ordered supramolecular complex between the dihydropyridine receptor (DHPR) and the ryanodine receptor (RyR1) mediates the quasi-instantaneous conversion from T-tubule depolarization into Ca 2+ release from the sarcoplasmic reticulum (SR). The DHPR has several key modules required for EC coupling: the voltage sensors and II-III loop in the alpha1s subunit, and the beta subunit. To gain insight into their molecular organization, this review examines the most updated 3D structure of the DHPR as obtained by transmission electron microscopy and image reconstruction. Although structure determination of a heteromeric membrane protein such as the DHPR is challenging, novel technical advances in protein expression and 3D labeling facilitated this task. The 3D structure of the DHPR complex consists of a main body with five irregular corners around its perimeter encompassing the transmembrane alpha 1s subunit besides the intracellular beta subunit, an extended extracellular alpha 2 subunit, and a bulky intracellular II-III loop. The structural definition attained at 19 Å resolution enabled docking of the atomic coordinates of structural homologs of the alpha1s and beta subunits. These structural features, together with their relative location with respect to the RyR1, are discussed in the context of the functional data.

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confidence: 74%

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confidence: 66%

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confidence: 99%

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confidence: 99%

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3D structure of muscle dihydropyridine receptor

Samsó¹

2015

Eur J Transl Myol

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“…1, top right). Ahead of the latter structure, Szpyt et al (2012) presented a lower resolution structure (∼19 Å) that identified many components of the Ca V 1.1 channel complex. The crystal structure of the Na V Ab bacterial Na + channel (Payandeh et al, 2011) was used to estimate the position and orientation of the α 1S subunit with the cryo-EM structure.…”

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confidence: 99%

Bridging the myoplasmic gap II: more recent advances in skeletal muscle excitation–contraction coupling

Bannister

2016

Journal of Experimental Biology

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In skeletal muscle, excitation-contraction (EC) coupling relies on the transmission of an intermolecular signal from the voltage-sensing regions of the L-type Ca 2+ channel (Ca V 1.1) in the plasma membrane to the channel pore of the type 1 ryanodine receptor (RyR1) nearly 10 nm away in the membrane of the sarcoplasmic reticulum (SR). Even though the roles of Ca V 1.1 and RyR1 as voltage sensor and SR Ca 2+ release channel, respectively, have been established for nearly 25 years, the mechanism underlying communication between these two channels remains undefined. In the course of this article, I will review current viewpoints on this topic with particular emphasis on recent studies.

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A guide to the 3D structure of the ryanodine receptor type 1 by cryoEM

Samsó

2016

Protein Science

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Signal transduction by the ryanodine receptor (RyR) is essential in many excitable cells including all striated contractile cells and some types of neurons. While its transmembrane domain is a classic tetrameric, six-transmembrane cation channel, the cytoplasmic domain is uniquely large and complex, hosting a multiplicity of specialized domains. The overall outline and substructure readily recognizable by electron microscopy make RyR a geometrically well-behaved specimen. Hence, for the last two decades, the 3D structural study of the RyR has tracked closely the technological advances in electron microscopy, cryo-electron microscopy (cryoEM), and computerized 3D reconstruction. This review summarizes the progress in the structural determination of RyR by cryoEM and, bearing in mind the leap in resolution provided by the recent implementation of direct electron detection, analyzes the first near-atomic structures of RyR. These reveal a complex orchestration of domains controlling the channel's function, and help to understand how this could break down as a consequence of disease-causing mutations.

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Three-Dimensional Localization of the α and β Subunits and of the II-III Loop in the Skeletal Muscle L-type Ca2+ Channel

Cited by 12 publications

References 71 publications

3D structure of muscle dihydropyridine receptor

3D structure of muscle dihydropyridine receptor

Bridging the myoplasmic gap II: more recent advances in skeletal muscle excitation–contraction coupling

A guide to the 3D structure of the ryanodine receptor type 1 by cryoEM

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