International Journal of Molecular Sciences

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15 pages, 3320 KiB

Open AccessArticle

Atomistic Structure and Dynamics of the Ca²⁺-ATPase Bound to Phosphorylated Phospholamban

by Rodrigo Aguayo-Ortiz and L. Michel Espinoza-Fonseca

Int. J. Mol. Sci. 2020, 21(19), 7261; https://doi.org/10.3390/ijms21197261 - 1 Oct 2020

Cited by 6 | Viewed by 2305

Abstract

Sarcoplasmic reticulum Ca²⁺-ATPase (SERCA) and phospholamban (PLB) are essential components of the cardiac Ca²⁺ transport machinery. PLB phosphorylation at residue Ser16 (pSer16) enhances SERCA activity in the heart via an unknown structural mechanism. Here, we report a fully atomistic model [...] Read more.

Sarcoplasmic reticulum Ca²⁺-ATPase (SERCA) and phospholamban (PLB) are essential components of the cardiac Ca²⁺ transport machinery. PLB phosphorylation at residue Ser16 (pSer16) enhances SERCA activity in the heart via an unknown structural mechanism. Here, we report a fully atomistic model of SERCA bound to phosphorylated PLB and study its structural dynamics on the microsecond time scale using all-atom molecular dynamics simulations in an explicit lipid bilayer and water environment. The unstructured N-terminal phosphorylation domain of PLB samples different orientations and covers a broad area of the cytosolic domain of SERCA but forms a stable complex mediated by pSer16 interactions with a binding site formed by SERCA residues Arg324/Lys328. PLB phosphorylation does not affect the interaction between the transmembrane regions of the two proteins; however, pSer16 stabilizes a disordered structure of the N-terminal phosphorylation domain that releases key inhibitory contacts between SERCA and PLB. We found that PLB phosphorylation is sufficient to guide the structural transitions of the cytosolic headpiece that are required to produce a competent structure of SERCA. We conclude that PLB phosphorylation serves as an allosteric molecular switch that releases inhibitory contacts and strings together the catalytic elements required for SERCA activation. This atomistic model represents a vivid atomic-resolution visualization of SERCA bound to phosphorylated PLB and provides previously inaccessible insights into the structural mechanism by which PLB phosphorylation releases SERCA inhibition in the heart. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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21 pages, 3570 KiB

Open AccessArticle

The Penta-EF-Hand ALG-2 Protein Interacts with the Cytosolic Domain of the SOCE Regulator SARAF and Interferes with Ubiquitination

by Wei Zhang, Ayaka Muramatsu, Rina Matsuo, Naoki Teranishi, Yui Kahara, Terunao Takahara, Hideki Shibata and Masatoshi Maki

Int. J. Mol. Sci. 2020, 21(17), 6315; https://doi.org/10.3390/ijms21176315 - 31 Aug 2020

Cited by 7 | Viewed by 3205

Abstract

ALG-2 is a penta-EF-hand Ca²⁺-binding protein and interacts with a variety of proteins in mammalian cells. In order to find new ALG-2-binding partners, we searched a human protein database and retrieved sequences containing the previously identified ALG-2-binding motif type 2 (ABM-2). [...] Read more.

ALG-2 is a penta-EF-hand Ca²⁺-binding protein and interacts with a variety of proteins in mammalian cells. In order to find new ALG-2-binding partners, we searched a human protein database and retrieved sequences containing the previously identified ALG-2-binding motif type 2 (ABM-2). After selecting 12 high-scored sequences, we expressed partial or full-length GFP-fused proteins in HEK293 cells and performed a semi-quantitative in vitro binding assay. SARAF, a negative regulator of store-operated Ca²⁺ entry (SOCE), showed the strongest binding activity. Biochemical analysis of Strep-tagged and GFP-fused SARAF proteins revealed ubiquitination that proceeded during pulldown assays under certain buffer conditions. Overexpression of ALG-2 interfered with ubiquitination of wild-type SARAF but not ubiquitination of the F228S mutant that had impaired ALG-2-binding activity. The SARAF cytosolic domain (CytD) contains two PPXY motifs targeted by the WW domains of NEDD4 family E3 ubiquitin ligases. The PPXY motif proximal to the ABM-2 sequence was found to be more important for both in-cell ubiquitination and post-cell lysis ubiquitination. A ubiquitination-defective mutant of SARAF with Lys-to-Arg substitutions in the CytD showed a slower degradation rate by half-life analysis. ALG-2 promoted Ca²⁺-dependent CytD-to-CytD interactions of SARAF. The ALG-2 dimer may modulate the stability of SARAF by sterically blocking ubiquitination and by bridging SARAF molecules at the CytDs. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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21 pages, 4979 KiB

Open AccessArticle

Knockout of stim2a Increases Calcium Oscillations in Neurons and Induces Hyperactive-Like Phenotype in Zebrafish Larvae

by Rishikesh Kumar Gupta, Iga Wasilewska, Oksana Palchevska and Jacek Kuźnicki

Int. J. Mol. Sci. 2020, 21(17), 6198; https://doi.org/10.3390/ijms21176198 - 27 Aug 2020

Cited by 10 | Viewed by 3192

Abstract

Stromal interaction molecule (STIM) proteins play a crucial role in store-operated calcium entry (SOCE) as endoplasmic reticulum Ca²⁺ sensors. In neurons, STIM2 was shown to have distinct functions from STIM1. However, its role in brain activity and behavior was not fully elucidated. [...] Read more.

Stromal interaction molecule (STIM) proteins play a crucial role in store-operated calcium entry (SOCE) as endoplasmic reticulum Ca²⁺ sensors. In neurons, STIM2 was shown to have distinct functions from STIM1. However, its role in brain activity and behavior was not fully elucidated. The present study analyzed behavior in zebrafish (Danio rerio) that lacked stim2a. The mutant animals had no morphological abnormalities and were fertile. RNA-sequencing revealed alterations of the expression of transcription factor genes and several members of the calcium toolkit. Neuronal Ca²⁺ activity was measured in vivo in neurons that expressed the GCaMP5G sensor. Optic tectum neurons in stim2a^−/− fish had more frequent Ca²⁺ signal oscillations compared with neurons in wildtype (WT) fish. We detected an increase in activity during the visual–motor response test, an increase in thigmotaxis in the open field test, and the disruption of phototaxis in the dark/light preference test in stim2a^−/− mutants compared with WT. Both groups of animals reacted to glutamate and pentylenetetrazol with an increase in activity during the visual–motor response test, with no major differences between groups. Altogether, our results suggest that the hyperactive-like phenotype of stim2a^−/− mutant zebrafish is caused by the dysregulation of Ca²⁺ homeostasis and signaling. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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28 pages, 5574 KiB

Open AccessArticle

Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

by Claes-Göran Reibring, Kristina Hallberg, Anders Linde and Amel Gritli-Linde

Int. J. Mol. Sci. 2020, 21(4), 1264; https://doi.org/10.3390/ijms21041264 - 13 Feb 2020

Cited by 7 | Viewed by 3535

Abstract

In mammals Homer1, Homer2 and Homer3 constitute a family of scaffolding proteins with key roles in Ca²⁺ signaling and Ca²⁺ transport. In rodents, Homer proteins and mRNAs have been shown to be expressed in various postnatal tissues and to be enriched [...] Read more.

In mammals Homer1, Homer2 and Homer3 constitute a family of scaffolding proteins with key roles in Ca²⁺ signaling and Ca²⁺ transport. In rodents, Homer proteins and mRNAs have been shown to be expressed in various postnatal tissues and to be enriched in brain. However, whether the Homers are expressed in developing tissues is hitherto largely unknown. In this work, we used immunohistochemistry and in situ hybridization to analyze the expression patterns of Homer1, Homer2 and Homer3 in developing cephalic structures. Our study revealed that the three Homer proteins and their encoding genes are expressed in a wide range of developing tissues and organs, including the brain, tooth, eye, cochlea, salivary glands, olfactory and respiratory mucosae, bone and taste buds. We show that although overall the three Homers exhibit overlapping distribution patterns, the proteins localize at distinct subcellular domains in several cell types, that in both undifferentiated and differentiated cells Homer proteins are concentrated in puncta and that the vascular endothelium is enriched with Homer3 mRNA and protein. Our findings suggest that Homer proteins may have differential and overlapping functions and are expected to be of value for future research aiming at deciphering the roles of Homer proteins during embryonic development. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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19 pages, 5369 KiB

Open AccessArticle

Constitutive Activation of Guanylate Cyclase by the G86R GCAP1 Variant Is Due to “Locking” Cation-π Interactions that Impair the Activator-to-Inhibitor Structural Transition

by Seher Abbas, Valerio Marino, Laura Bielefeld, Karl-Wilhelm Koch and Daniele Dell’Orco

Int. J. Mol. Sci. 2020, 21(3), 752; https://doi.org/10.3390/ijms21030752 - 23 Jan 2020

Cited by 7 | Viewed by 2982

Abstract

Guanylate Cyclase activating protein 1 (GCAP1) mediates the Ca²⁺-dependent regulation of the retinal Guanylate Cyclase (GC) in photoreceptors, acting as a target inhibitor at high [Ca²⁺] and as an activator at low [Ca²⁺]. Recently, a novel missense [...] Read more.

Guanylate Cyclase activating protein 1 (GCAP1) mediates the Ca²⁺-dependent regulation of the retinal Guanylate Cyclase (GC) in photoreceptors, acting as a target inhibitor at high [Ca²⁺] and as an activator at low [Ca²⁺]. Recently, a novel missense mutation (G86R) was found in GUCA1A, the gene encoding for GCAP1, in patients diagnosed with cone-rod dystrophy. The G86R substitution was found to affect the flexibility of the hinge region connecting the N- and C-domains of GCAP1, resulting in decreased Ca^2+-sensitivity and abnormally enhanced affinity for GC. Based on a structural model of GCAP1, here, we tested the hypothesis of a cation-π interaction between the positively charged R86 and the aromatic W94 as the main mechanism underlying the impaired activator-to-inhibitor conformational change. W94 was mutated to F or L, thus, resulting in the double mutants G86R+W94L/F. The double mutants showed minor structural and stability changes with respect to the single G86R mutant, as well as lower affinity for both Mg²⁺ and Ca²⁺, moreover, substitutions of W94 abolished “phase II” in Ca²⁺-titrations followed by intrinsic fluorescence. Interestingly, the presence of an aromatic residue in position 94 significantly increased the aggregation propensity of Ca²⁺-loaded GCAP1 variants. Finally, atomistic simulations of all GCAP1 variants in the presence of Ca²⁺ supported the presence of two cation-π interactions involving R86, which was found to act as a bridge between W94 and W21, thus, locking the hinge region in an activator-like conformation and resulting in the constitutive activation of the target under physiological conditions. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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18 pages, 1850 KiB

Open AccessArticle

Methylene Blue Blocks and Reverses the Inhibitory Effect of Tau on PMCA Function

by Maria Berrocal, Montaña Caballero-Bermejo, Carlos Gutierrez-Merino and Ana M. Mata

Int. J. Mol. Sci. 2019, 20(14), 3521; https://doi.org/10.3390/ijms20143521 - 18 Jul 2019

Cited by 14 | Viewed by 5526

Abstract

Methylene blue (MB) is a synthetic phenothiazine dye that, in the last years, has generated much debate about whether it could be a useful therapeutic drug for tau-related pathologies, such as Alzheimer’s disease (AD). However, the molecular mechanism of action is far from [...] Read more.

Methylene blue (MB) is a synthetic phenothiazine dye that, in the last years, has generated much debate about whether it could be a useful therapeutic drug for tau-related pathologies, such as Alzheimer’s disease (AD). However, the molecular mechanism of action is far from clear. Recently we reported that MB activates the plasma membrane Ca²⁺-ATPase (PMCA) in membranes from human and pig tissues and from cells cultures, and that it could protect against inactivation of PMCA by amyloid β-peptide (Aβ). The purpose of the present study is to further examine whether the MB could also modulate the inhibitory effect of tau, another key molecular marker of AD, on PMCA activity. By using kinetic assays in membranes from several tissues and cell cultures, we found that this phenothiazine was able to block and even to completely reverse the inhibitory effect of tau on PMCA. The results of this work point out that MB could mediate the toxic effect of tau related to the deregulation of calcium homeostasis by blocking the impairment of PMCA activity by tau. We then could conclude that MB could interfere with the toxic effects of tau by restoring the function of PMCA pump as a fine tuner of calcium homeostasis. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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22 pages, 5606 KiB

Open AccessArticle

Changes of Thermostability, Organic Solvent, and pH Stability in Geobacillus zalihae HT1 and Its Mutant by Calcium Ion

by Siti Nor Hasmah Ishak, Malihe Masomian, Nor Hafizah Ahmad Kamarudin, Mohd Shukuri Mohamad Ali, Thean Chor Leow and Raja Noor Zaliha Raja Abd. Rahman

Int. J. Mol. Sci. 2019, 20(10), 2561; https://doi.org/10.3390/ijms20102561 - 24 May 2019

Cited by 24 | Viewed by 5087

Abstract

Thermostable T1 lipase from Geobacillus zalihae has been crystallized using counter-diffusion method under space and Earth conditions. The comparison of the three-dimensional structures from both crystallized proteins show differences in the formation of hydrogen bond and ion interactions. Hydrogen bond and ion interaction [...] Read more.

Thermostable T1 lipase from Geobacillus zalihae has been crystallized using counter-diffusion method under space and Earth conditions. The comparison of the three-dimensional structures from both crystallized proteins show differences in the formation of hydrogen bond and ion interactions. Hydrogen bond and ion interaction are important in the stabilization of protein structure towards extreme temperature and organic solvents. In this study, the differences of hydrogen bond interactions at position Asp43, Thr118, Glu250, and Asn304 and ion interaction at position Glu226 was chosen to imitate space-grown crystal structure, and the impact of these combined interactions in T1 lipase-mutated structure was studied. Using space-grown T1 lipase structure as a reference, subsequent simultaneous mutation D43E, T118N, E226D, E250L, and N304E was performed on recombinant wild-type T1 lipase (wt-HT1) to generate a quintuple mutant term as 5M mutant lipase. This mutant lipase shared similar characteristics to its wild-type in terms of optimal pH and temperature. The stability of mutant 5M lipase improved significantly in acidic and alkaline pH as compared to wt-HT1. 5M lipase was highly stable in organic solvents such as dimethyl sulfoxide (DMSO), methanol, and n-hexane compared to wt-HT1. Both wild-type and mutant lipases were found highly activated in calcium as compared to other metal ions due to the presence of calcium-binding site for thermostability. The presence of calcium prolonged the half-life of mutant 5M and wt-HT1, and at the same time increased their melting temperature (T_m). The melting temperature of 5M and wt-HT1 lipases increased at 8.4 and 12.1 °C, respectively, in the presence of calcium as compared to those without. Calcium enhanced the stability of mutant 5M in 25% (v/v) DMSO, n-hexane, and n-heptane. The lipase activity of wt-HT1 also increased in 25% (v/v) ethanol, methanol, acetonitrile, n-hexane, and n-heptane in the presence of calcium. The current study showed that the accumulation of amino acid substitutions D43E, T118N, E226D, E250L, and N304E produced highly stable T1 mutant when hydrolyzing oil in selected organic solvents such as DMSO, n-hexane, and n-heptane. It is also believed that calcium ion plays important role in regulating lipase thermostability. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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12 pages, 2080 KiB

Open AccessArticle

Inhibition of Calcium/Calmodulin-Dependent Protein Kinase Kinase β Is Detrimental in Hypoxia–Ischemia Neonatal Brain Injury

by Jia-Wei Min, Fan Bu, Li Qi, Yashasvee Munshi, Gab Seok Kim, Sean P. Marrelli, Louise D. McCullough and Jun Li

Int. J. Mol. Sci. 2019, 20(9), 2063; https://doi.org/10.3390/ijms20092063 - 26 Apr 2019

Cited by 6 | Viewed by 3803

Abstract

Neonatal hypoxia–ischemia (HI) is a major cause of death and disability in neonates. HI leads to a dramatic rise in intracellular calcium levels, which was originally thought to be detrimental to the brain. However, it has been increasingly recognized that this calcium signaling [...] Read more.

Neonatal hypoxia–ischemia (HI) is a major cause of death and disability in neonates. HI leads to a dramatic rise in intracellular calcium levels, which was originally thought to be detrimental to the brain. However, it has been increasingly recognized that this calcium signaling may also play an important protective role after injury by triggering endogenous neuroprotective pathways. Calcium/calmodulin-dependent protein kinase kinase β (CaMKK β) is a major kinase activated by elevated levels of intracellular calcium. Here we evaluated the functional role of CaMKK β in neonatal mice after HI in both acute and chronic survival experiments. Postnatal day ten wild-type (WT) and CaMKK β knockout (KO) mouse male pups were subjected to unilateral carotid artery ligation, followed by 40 min of hypoxia (10% O₂ in N₂). STO-609, a CaMKK inhibitor, was administered intraperitoneally to WT mice at 5 minutes after HI. TTC (2,3,5-triphenyltetrazolium chloride monohydrate) staining was used to assess infarct volume 24 h after HI. CaMKK β KO mice had larger infarct volume than WT mice and STO-609 increased the infarct volume in WT mice after HI. In chronic survival experiments, WT mice treated with STO-609 showed increased tissue loss in the ipsilateral hemisphere three weeks after HI. Furthermore, when compared with vehicle-treated mice, they showed poorer functional recovery during the three week survival period, as measured by the wire hang test and corner test. Loss of blood–brain barrier proteins, a reduction in survival protein (Bcl-2), and an increase in pro-apoptotic protein Bax were also seen after HI with CaMKK β inhibition. In conclusion, inhibition of CaMKK β exacerbated neonatal hypoxia–ischemia injury in mice. Our data suggests that enhancing CaMKK signaling could be a potential target for the treatment of hypoxic–ischemic brain injury. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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15 pages, 2017 KiB

Open AccessArticle

Metabolic Inhibition Induces Transient Increase of L-type Ca²⁺ Current in Human and Rat Cardiac Myocytes

by Rimantas Treinys, Giedrius Kanaporis, Rodolphe Fischmeister and Jonas Jurevičius

Int. J. Mol. Sci. 2019, 20(6), 1501; https://doi.org/10.3390/ijms20061501 - 26 Mar 2019

Cited by 7 | Viewed by 4110

Abstract

Metabolic inhibition is a common condition observed during ischemic heart disease and heart failure. It is usually accompanied by a reduction in L-type Ca²⁺ channel (LTCC) activity. In this study, however, we show that metabolic inhibition results in a biphasic effect on [...] Read more.

Metabolic inhibition is a common condition observed during ischemic heart disease and heart failure. It is usually accompanied by a reduction in L-type Ca²⁺ channel (LTCC) activity. In this study, however, we show that metabolic inhibition results in a biphasic effect on LTCC current (I_CaL) in human and rat cardiac myocytes: an initial increase of I_CaL is observed in the early phase of metabolic inhibition which is followed by the more classical and strong inhibition. We studied the mechanism of the initial increase of I_CaL in cardiac myocytes during β-adrenergic stimulation by isoprenaline, a non-selective agonist of β-adrenergic receptors. The whole-cell patch–clamp technique was used to record the I_CaL in single cardiac myocytes. The initial increase of I_CaL was induced by a wide range of metabolic inhibitors (FCCP, 2,4-DNP, rotenone, antimycin A). In rat cardiomyocytes, the initial increase of I_CaL was eliminated when the cells were pre-treated with thapsigargin leading to the depletion of Ca²⁺ from the sarcoplasmic reticulum (SR). Similar results were obtained when Ca²⁺ release from the SR was blocked with ryanodine. These data suggest that the increase of I_CaL in the early phase of metabolic inhibition is due to a reduced calcium dependent inactivation (CDI) of LTCCs. This was further confirmed in human atrial myocytes where FCCP failed to induce the initial stimulation of I_CaL when Ca²⁺ was replaced by Ba²⁺, eliminating CDI of LTCCs. We conclude that the initial increase in I_CaL observed during the metabolic inhibition in human and rat cardiomyocytes is a consequence of an acute reduction of Ca²⁺ release from SR resulting in reduced CDI of LTCCs. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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17 pages, 4525 KiB

Open AccessArticle

Inhibition of Triple-Negative Breast Cancer Cell Aggressiveness by Cathepsin D Blockage: Role of Annexin A1

by Mariana Alves Pereira Zóia, Fernanda Van Petten Azevedo, Lara Vecchi, Sara Teixeira Soares Mota, Vinícius de Rezende Rodovalho, Antonielle Oliveira Cordeiro, Lucas Ian Veloso Correia, Anielle Christine Almeida Silva, Veridiana de Melo Rodrigues Ávila, Thaise Gonçalves de Araújo and Luiz Ricardo Goulart

Int. J. Mol. Sci. 2019, 20(6), 1337; https://doi.org/10.3390/ijms20061337 - 16 Mar 2019

Cited by 17 | Viewed by 4406

Abstract

Triple-negative breast cancers (TNBCs) are more aggressive than other breast cancer (BC) subtypes and lack effective therapeutic options. Unraveling marker events of TNBCs may provide new directions for development of strategies for targeted TNBC therapy. Herein, we reported that Annexin A1 (AnxA1) and [...] Read more.

Triple-negative breast cancers (TNBCs) are more aggressive than other breast cancer (BC) subtypes and lack effective therapeutic options. Unraveling marker events of TNBCs may provide new directions for development of strategies for targeted TNBC therapy. Herein, we reported that Annexin A1 (AnxA1) and Cathepsin D (CatD) are highly expressed in MDA-MB-231 (TNBC lineage), compared to MCF-10A and MCF-7. Since the proposed concept was that CatD has protumorigenic activity associated with its ability to cleave AnxA1 (generating a 35.5 KDa fragment), we investigated this mechanism more deeply using the inhibitor of CatD, Pepstatin A (PepA). Fourier Transform Infrared (FTIR) spectroscopy demonstrated that PepA inhibits CatD activity by occupying its active site; the OH bond from PepA interacts with a CO bond from carboxylic acids of CatD catalytic aspartate dyad, favoring the deprotonation of Asp³³ and consequently inhibiting CatD. Treatment of MDA-MB-231 cells with PepA induced apoptosis and autophagy processes while reducing the proliferation, invasion, and migration. Finally, in silico molecular docking demonstrated that the catalytic inhibition comprises Asp²³¹ protonated and Asp³³ deprotonated, proving all functional results obtained. Our findings elucidated critical CatD activity in TNBC cell trough AnxA1 cleavage, indicating the inhibition of CatD as a possible strategy for TNBC treatment. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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Review

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20 pages, 1457 KiB

Open AccessReview

ASGR1 and Its Enigmatic Relative, CLEC10A

by J. Kenneth Hoober

Int. J. Mol. Sci. 2020, 21(14), 4818; https://doi.org/10.3390/ijms21144818 - 8 Jul 2020

Cited by 32 | Viewed by 6859

Abstract

The large family of C-type lectin (CLEC) receptors comprises carbohydrate-binding proteins that require Ca²⁺ to bind a ligand. The prototypic receptor is the asialoglycoprotein receptor-1 (ASGR1, CLEC4H1) that is expressed primarily by hepatocytes. The early work on ASGR1, which is highly specific [...] Read more.

The large family of C-type lectin (CLEC) receptors comprises carbohydrate-binding proteins that require Ca²⁺ to bind a ligand. The prototypic receptor is the asialoglycoprotein receptor-1 (ASGR1, CLEC4H1) that is expressed primarily by hepatocytes. The early work on ASGR1, which is highly specific for N-acetylgalactosamine (GalNAc), established the foundation for understanding the overall function of CLEC receptors. Cells of the immune system generally express more than one CLEC receptor that serve diverse functions such as pathogen-recognition, initiation of cellular signaling, cellular adhesion, glycoprotein turnover, inflammation and immune responses. The receptor CLEC10A (C-type lectin domain family 10 member A, CD301; also called the macrophage galactose-type lectin, MGL) contains a carbohydrate-recognition domain (CRD) that is homologous to the CRD of ASGR1, and thus, is also specific for GalNAc. CLEC10A is most highly expressed on immature DCs, monocyte-derived DCs, and alternatively activated macrophages (subtype M2a) as well as oocytes and progenitor cells at several stages of embryonic development. This receptor is involved in initiation of T_H1, T_H2, and T_H17 immune responses and induction of tolerance in naïve T cells. Ligand-mediated endocytosis of CLEC receptors initiates a Ca²⁺ signal that interestingly has different outcomes depending on ligand properties, concentration, and frequency of administration. This review summarizes studies that have been carried out on these receptors. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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32 pages, 5030 KiB

Open AccessReview

Linking Biochemical and Structural States of SERCA: Achievements, Challenges, and New Opportunities

by Rodrigo Aguayo-Ortiz and L. Michel Espinoza-Fonseca

Int. J. Mol. Sci. 2020, 21(11), 4146; https://doi.org/10.3390/ijms21114146 - 10 Jun 2020

Cited by 39 | Viewed by 7271

Abstract

Sarcoendoplasmic reticulum calcium ATPase (SERCA), a member of the P-type ATPase family of ion and lipid pumps, is responsible for the active transport of Ca²⁺ from the cytoplasm into the sarcoplasmic reticulum lumen of muscle cells, into the endoplasmic reticulum (ER) of [...] Read more.

Sarcoendoplasmic reticulum calcium ATPase (SERCA), a member of the P-type ATPase family of ion and lipid pumps, is responsible for the active transport of Ca²⁺ from the cytoplasm into the sarcoplasmic reticulum lumen of muscle cells, into the endoplasmic reticulum (ER) of non-muscle cells. X-ray crystallography has proven to be an invaluable tool in understanding the structural changes of SERCA, and more than 70 SERCA crystal structures representing major biochemical states (defined by bound ligand) have been deposited in the Protein Data Bank. Consequently, SERCA is one of the best characterized components of the calcium transport machinery in the cell. Emerging approaches in the field, including spectroscopy and molecular simulation, now help integrate and interpret this rich structural information to understand the conformational transitions of SERCA that occur during activation, inhibition, and regulation. In this review, we provide an overview of the crystal structures of SERCA, focusing on identifying metrics that facilitate structure-based categorization of major steps along the catalytic cycle. We examine the integration of crystallographic data with different biophysical approaches and computational methods to link biochemical and structural states of SERCA that are populated in the cell. Finally, we discuss the challenges and new opportunities in the field, including structural elucidation of functionally important and novel regulatory complexes of SERCA, understanding the structural basis of functional divergence among homologous SERCA regulators, and bridging the gap between basic and translational research directed toward therapeutic modulation of SERCA. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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19 pages, 1610 KiB

Open AccessReview

Intracellular Ca²⁺ Signalling in the Pathogenesis of Acute Pancreatitis: Recent Advances and Translational Perspectives

by Petra Pallagi, Tamara Madácsy, Árpád Varga and József Maléth

Int. J. Mol. Sci. 2020, 21(11), 4005; https://doi.org/10.3390/ijms21114005 - 3 Jun 2020

Cited by 50 | Viewed by 8522

Abstract

Intracellular Ca²⁺ signalling is a major signal transductional pathway in non-excitable cells, responsible for the regulation of a variety of physiological functions. In the secretory epithelial cells of the exocrine pancreas, such as acinar and ductal cells, intracellular Ca²⁺ elevation regulates [...] Read more.

Intracellular Ca²⁺ signalling is a major signal transductional pathway in non-excitable cells, responsible for the regulation of a variety of physiological functions. In the secretory epithelial cells of the exocrine pancreas, such as acinar and ductal cells, intracellular Ca²⁺ elevation regulates digestive enzyme secretion in acini or fluid and ion secretion in ductal cells. Although Ca²⁺ is a uniquely versatile orchestrator of epithelial physiology, unregulated global elevation of the intracellular Ca²⁺ concentration is an early trigger for the development of acute pancreatitis (AP). Regardless of the aetiology, different forms of AP all exhibit sustained intracellular Ca²⁺ elevation as a common hallmark. The release of endoplasmic reticulum (ER) Ca²⁺ stores by toxins (such as bile acids or fatty acid ethyl esters (FAEEs)) or increased intrapancreatic pressure activates the influx of extracellular Ca²⁺ via the Orai1 Ca²⁺ channel, a process known as store-operated Ca²⁺ entry (SOCE). Intracellular Ca²⁺ overload can lead to premature activation of trypsinogen in pancreatic acinar cells and impaired fluid and HCO₃^- secretion in ductal cells. Increased and unbalanced reactive oxygen species (ROS) production caused by sustained Ca²⁺ elevation further contributes to cell dysfunction, leading to mitochondrial damage and cell death. Translational studies of AP identified several potential target molecules that can be modified to prevent intracellular Ca²⁺ overload. One of the most promising drugs, a selective inhibitor of the Orai1 channel that has been shown to inhibit extracellular Ca²⁺ influx and protect cells from injury, is currently being tested in clinical trials. In this review, we will summarise the recent advances in the field, with a special focus on the translational aspects of the basic findings. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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13 pages, 981 KiB

Open AccessReview

Valosin-Containing Protein, a Calcium-Associated ATPase Protein, in Endoplasmic Reticulum and Mitochondrial Function and Its Implications for Diseases

by Xiaonan Sun and Hongyu Qiu

Int. J. Mol. Sci. 2020, 21(11), 3842; https://doi.org/10.3390/ijms21113842 - 28 May 2020

Cited by 32 | Viewed by 4654

Abstract

Endoplasmic reticulum (ER) and mitochondrion are the key organelles in mammal cells and play crucial roles in a variety of biological functions in both physiological and pathological conditions. Valosin-containing protein (VCP), a newly identified calcium-associated ATPase protein, has been found to be involved [...] Read more.

Endoplasmic reticulum (ER) and mitochondrion are the key organelles in mammal cells and play crucial roles in a variety of biological functions in both physiological and pathological conditions. Valosin-containing protein (VCP), a newly identified calcium-associated ATPase protein, has been found to be involved in both ER and mitochondrial function. Impairment of VCP, caused by structural mutations or alterations of expressions, contributes to the development of various diseases, through an integrating effect on ER, mitochondria and the ubiquitin–proteasome system, by interfering with protein degradation, subcellular translocation and calcium homeostasis. Thus, understanding the role and the molecular mechanisms of VCP in these organelles brings new insights to the pathogenesis of the associated diseases, and leads to the discovery of new therapeutic strategies. In this review, we summarized the progress of studies on VCP, in terms of its regulation of ER and mitochondrial function and its implications for the associated diseases, focusing on the cancers, heart disease, and neurodegenerative disorders. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling 2.0)

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22 pages, 2740 KiB

Open AccessReview

Calcium Mechanisms in Limb-Girdle Muscular Dystrophy with CAPN3 Mutations

by Jaione Lasa-Elgarresta, Laura Mosqueira-Martín, Neia Naldaiz-Gastesi, Amets Sáenz, Adolfo López de Munain and Ainara Vallejo-Illarramendi

Int. J. Mol. Sci. 2019, 20(18), 4548; https://doi.org/10.3390/ijms20184548 - 13 Sep 2019

Cited by 27 | Viewed by 7235

Abstract

Limb-girdle muscular dystrophy recessive 1 (LGMDR1), previously known as LGMD2A, is a rare disease caused by mutations in the CAPN3 gene. It is characterized by progressive weakness of shoulder, pelvic, and proximal limb muscles that usually appears in children and young adults and [...] Read more.

Limb-girdle muscular dystrophy recessive 1 (LGMDR1), previously known as LGMD2A, is a rare disease caused by mutations in the CAPN3 gene. It is characterized by progressive weakness of shoulder, pelvic, and proximal limb muscles that usually appears in children and young adults and results in loss of ambulation within 20 years after disease onset in most patients. The pathophysiological mechanisms involved in LGMDR1 remain mostly unknown, and to date, there is no effective treatment for this disease. Here, we review clinical and experimental evidence suggesting that dysregulation of Ca²⁺ homeostasis in the skeletal muscle is a significant underlying event in this muscular dystrophy. We also review and discuss specific clinical features of LGMDR1, CAPN3 functions, novel putative targets for therapeutic strategies, and current approaches aiming to treat LGMDR1. These novel approaches may be clinically relevant not only for LGMDR1 but also for other muscular dystrophies with secondary calpainopathy or with abnormal Ca²⁺ homeostasis, such as LGMD2B/LGMDR2 or sporadic inclusion body myositis. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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28 pages, 2069 KiB

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Calcium-Binding Proteins in the Nervous System during Hibernation: Neuroprotective Strategies in Hypometabolic Conditions?

by Giacomo Gattoni and Graziella Bernocchi

Int. J. Mol. Sci. 2019, 20(9), 2364; https://doi.org/10.3390/ijms20092364 - 13 May 2019

Cited by 12 | Viewed by 5496

Abstract

Calcium-binding proteins (CBPs) can influence and react to Ca²⁺ transients and modulate the activity of proteins involved in both maintaining homeostatic conditions and protecting cells in harsh environmental conditions. Hibernation is a strategy that evolved in vertebrate and invertebrate species to survive [...] Read more.

Calcium-binding proteins (CBPs) can influence and react to Ca²⁺ transients and modulate the activity of proteins involved in both maintaining homeostatic conditions and protecting cells in harsh environmental conditions. Hibernation is a strategy that evolved in vertebrate and invertebrate species to survive in cold environments; it relies on molecular, cellular, and behavioral adaptations guided by the neuroendocrine system that together ensure unmatched tolerance to hypothermia, hypometabolism, and hypoxia. Therefore, hibernation is a useful model to study molecular neuroprotective adaptations to extreme conditions, and can reveal useful applications to human pathological conditions. In this review, we describe the known changes in Ca²⁺-signaling and the detection and activity of CBPs in the nervous system of vertebrate and invertebrate models during hibernation, focusing on cytosolic Ca²⁺ buffers and calmodulin. Then, we discuss these findings in the context of the neuroprotective and neural plasticity mechanisms in the central nervous system: in particular, those associated with cytoskeletal proteins. Finally, we compare the expression of CBPs in the hibernating nervous system with two different conditions of neurodegeneration, i.e., platinum-induced neurotoxicity and Alzheimer’s disease, to highlight the similarities and differences and demonstrate the potential of hibernation to shed light into part of the molecular mechanisms behind neurodegenerative diseases. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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22 pages, 1577 KiB

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Expression of Ca²⁺-Binding Buffer Proteins in the Human and Mouse Retinal Neurons

by Tamás Kovács-Öller, Gergely Szarka, Alma Ganczer, Ádám Tengölics, Boglárka Balogh and Béla Völgyi

Int. J. Mol. Sci. 2019, 20(9), 2229; https://doi.org/10.3390/ijms20092229 - 7 May 2019

Cited by 18 | Viewed by 4613

Abstract

Ca²⁺-binding buffer proteins (CaBPs) are widely expressed by various neurons throughout the central nervous system (CNS), including the retina. While the expression of CaBPs by photoreceptors, retinal interneurons and the output ganglion cells in the mammalian retina has been extensively studied, [...] Read more.

Ca²⁺-binding buffer proteins (CaBPs) are widely expressed by various neurons throughout the central nervous system (CNS), including the retina. While the expression of CaBPs by photoreceptors, retinal interneurons and the output ganglion cells in the mammalian retina has been extensively studied, a general description is still missing due to the differences between species, developmental expression patterns and study-to-study discrepancies. Furthermore, CaBPs are occasionally located in a compartment-specific manner and two or more CaBPs can be expressed by the same neuron, thereby sharing the labor of Ca²⁺ buffering in the intracellular milieu. This article reviews this topic by providing a framework on CaBP functional expression by neurons of the mammalian retina with an emphasis on human and mouse retinas and the three most abundant and extensively studied buffer proteins: parvalbumin, calretinin and calbindin. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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14 pages, 813 KiB

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Calcium-Binding Proteins as Determinants of Central Nervous System Neuronal Vulnerability to Disease

by Richard Fairless, Sarah K. Williams and Ricarda Diem

Int. J. Mol. Sci. 2019, 20(9), 2146; https://doi.org/10.3390/ijms20092146 - 30 Apr 2019

Cited by 73 | Viewed by 5912

Abstract

Neuronal subpopulations display differential vulnerabilities to disease, but the factors that determine their susceptibility are poorly understood. Toxic increases in intracellular calcium are a key factor in several neurodegenerative processes, with calcium-binding proteins providing an important first line of defense through their ability [...] Read more.

Neuronal subpopulations display differential vulnerabilities to disease, but the factors that determine their susceptibility are poorly understood. Toxic increases in intracellular calcium are a key factor in several neurodegenerative processes, with calcium-binding proteins providing an important first line of defense through their ability to buffer incoming calcium, allowing the neuron to quickly achieve homeostasis. Since neurons expressing different calcium-binding proteins have been reported to be differentially susceptible to degeneration, it can be hypothesized that rather than just serving as markers of different neuronal subpopulations, they might actually be a key determinant of survival. In this review, we will summarize some of the evidence that expression of the EF-hand calcium-binding proteins, calbindin, calretinin and parvalbumin, may influence the susceptibility of distinct neuronal subpopulations to disease processes. Full article

(This article belongs to the Special Issue Calcium-Binding Proteins and Cell Signaling)

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