Essential Ca
 2+
 -Binding Motif for Ca
 2+
 -Sensitive Inactivation of L-Type Ca
 2+
 Channels

Leon, Marita de; Wang, Yan; Jones, Lisa P.; Perez‐Reyes, Edward; Wei, Xing; Soong, Tuck Wah; Snutch, Terry P.; Yue, David T.

doi:10.1126/science.270.5241.1502

Cited by 261 publications

(219 citation statements)

References 36 publications

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“…34,46 Such a direct binding of Ca 2ϩ ions has been proposed by Gutnick et al 17 for molluscan Ca 2ϩ channels. de Leon et al 11 and Imredy and Yue 23 have recently provided evidence for such a direct effect on native and artificially expressed cardiac L-type Ca 2ϩ channels, and similar mechanisms may also exist in neuronal Land N-type Ca 2ϩ channels. In summary, our results provide strong evidence against the involvement of CaN in the inactivation process of L-and N-type Ca 2ϩ channels, and support the dominant role of direct binding of Ca 2ϩ ions to the channels.…”

Section: Discussionmentioning

confidence: 96%

Calcium-dependent inactivation of neuronal calcium channel currents is independent of calcineurin

Zeilhofer¹,

Blank

Neuhuber

et al. 1999

Neuroscience

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Abstract-Dephosphorylation by the Ca 2ϩ /calmodulin-dependent phosphatase calcineurin has been suggested as an important mechanism of Ca 2ϩ -dependent inactivation of voltage-gated Ca 2ϩ channels. We have tested whether calcineurin plays a role in the inactivation process of two types of high-voltage-activated Ca 2ϩ channels (L and N type) widely expressed in the central nervous system, using the immunosuppressive drug FK506 (tacrolimus), which inhibits calcineurin after binding to intracellular FK506 binding proteins. Inactivation of L-and N-type Ca 2ϩ channels was studied in a rat pituitary tumor cell line (GH 3 ) and chicken dorsal root ganglion neurons, respectively. With the use of antisera directed against the calcineurin subunit B and the 12,000 mol. wt binding protein, we show that both proteins are present in the cytoplasm of GH 3 cells and chicken dorsal root ganglion neurons. Ionic currents through voltage-gated Ca 2ϩ channels were investigated in the perforated-patch and whole-cell configurations of the patch-clamp technique. The inactivation of L-as well as N-type Ca 2ϩ currents could be well fitted with a bi-exponential function. Inactivation was largely reduced when Ba 2ϩ substituted for extracellular Ca 2ϩ or when the Ca 2ϩ chelator EGTA was present intracellularly, indicating that both types of Ca 2ϩ currents exhibited Ca 2ϩ -dependent inactivation. Extracellular (perforated-patch configuration) or intracellular (whole-cell configuration) application of FK506 to inactivate calcineurin had no effect on the amplitude and time-course of Ca 2ϩ channel current inactivation of either L-or N-type Ca 2ϩ channels. In addition, we found that recovery from inactivation and rundown of N-type Ca 2ϩ channel currents were not affected by FK506.Our results provide direct evidence that the calcium-dependent enzyme calcineurin is not involved in the inactivation process of the two Ca 2ϩ channel types which are important for neuronal functioning, such as gene expression and transmitter release. ᭧ 1999 IBRO. Published by Elsevier Science Ltd.Key words: Ca 2ϩ channel, inactivation, calcineurin, FK506, FKBP.An increase in the intracellular concentration of free Ca 2ϩ ([Ca 2ϩ ] i ) plays a pivotal role in a large variety of physiological functions, such as excitation-secretion coupling and neuronal plasticity. On the other hand, excessive increases in [Ca 2ϩ ] i may lead to excitotoxic cell death, which occurs, for example, during brain ischemia and epileptic seizures. 16 In neurons, one important pathway that leads to an increase in [Ca 2ϩ ] i is the opening of voltage-gated Ca 2ϩ channels in the plasma membrane. The activity of these Ca 2ϩ channels is effectively regulated by G-protein-coupled receptors and by Ca 2ϩ -and voltage-dependent inactivation of these ion channels. 3,13,40 Among the different types of Ca 2ϩ channels, Ca 2ϩ -dependent inactivation is best documented for L-type Ca 2ϩ channels. 2,23,24,34,46 However, a Ca 2ϩ -mediated component of Ca 2ϩ channel inactivation has also been described for N-typ...

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Section: Discussionmentioning

confidence: 96%

Calcium-dependent inactivation of neuronal calcium channel currents is independent of calcineurin

Zeilhofer¹,

Blank

Neuhuber

et al. 1999

Neuroscience

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“…18 It was hypothesized that CALP2, as a CaM antagonist, opens calcium channels in airway epithelial cells, leading to calcium influx in these cells. 18,53,54 Therefore, we now postulate that the production of radicals by CALP2 in macrophages is predominantly due to an increase of [Ca 2 þ ] i . Although less than in the presence of extracellular calcium, we found that macrophages still produce radicals upon stimulation with CALP2 in calcium-free medium.…”

Section: Discussionmentioning

confidence: 99%

Specific modulation of calmodulin activity induces a dramatic production of superoxide by alveolar macrophages

Broeke

Leusink-Muis

Hilberdink

et al. 2004

Laboratory Investigation

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Airway inflammation is a characteristic feature in airway diseases such as asthma and chronic obstructive pulmonary disease. Oxidative stress, caused by the excessive production of reactive oxygen species by inflammatory cells like macrophages, eosinophils and neutrophils, is thought to be important in the complex pathogenesis of such airway diseases. The calcium-sensing regulatory protein calmodulin (CaM) binds and regulates different target enzymes and proteins, including calcium channels. In the present study, we investigated whether CaM, via the modulation of calcium channel function, influences [Ca 2 þ ] i in pulmonary inflammatory cells, and consequently, modulates the production of reactive oxygen species by these cells. This was tested with a peptide termed calcium-like peptide 2 (CALP2), which was previously shown to regulate such channels. Specifically, radical production by purified broncho-alveolar lavage cells from guinea-pigs in response to CALP2 was measured. CALP2 was a strong activator of alveolar macrophages. In contrast, CALP2 was only a mild activator of neutrophils and did not induce radical production by eosinophils. The CALP2-induced radical production was mainly intracellular, and was completely blocked by the NADPH-oxidase inhibitor DPI, the superoxide inhibitor SOD and the CaM antagonist W7. Furthermore, the calcium channel blocker lanthanum partly inhibited the cellular activation by CALP2. We conclude that alveolar macrophages, but not neutrophils or eosinophils, can produce extremely high amounts of reactive oxygen species when stimulated via the calcium/CaM pathway. These results may contribute to new therapeutic strategies against oxidative stress in airway diseases.

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“…Initially, it was hypothesized that the Ca 2ϩ binding to the EF hand in PC-2 (amino acids 754 -782), as well as in the structurally related PCL and ␣ subunit of voltage-activated Ca 2ϩ channels, was the mediator of the Ca 2ϩ -dependent modulation of channel activity (2,33,34). Although studies have not directly addressed this in PC-2, in both polycystin-␣-like and the ␣ 1c voltage-activated Ca 2ϩ channel, this hypothesis has not been entirely borne out (33,35).…”

Section: Discussionmentioning

confidence: 99%

Calcium Dependence of Polycystin-2 Channel Activity Is Modulated by Phosphorylation at Ser812

Cai

Anyatonwu

Okuhara

et al. 2004

Journal of Biological Chemistry

140

188

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Polycystin-2 (PC-2) is a non-selective cation channel that, when mutated, results in autosomal dominant polycystic kidney disease. In an effort to understand the regulation of this channel, we investigated the role of protein phosphorylation in PC-2 function. We demonstrated the direct incorporation of phosphate into PC-2 in cells and tissues and found that this constitutive phosphorylation occurs at Ser 812 , a putative casein kinase II (CK2) substrate domain. Ser 812 can be phosphorylated by CK2 in vitro and substitution S812A results in failure to incorporate phosphate in cultured epithelial cells. Non-phosphorylated forms of PC-2 traffic normally in the endoplasmic reticulum and cilial compartments and retain homo-and hetero-multimerization interactions with PC-2 and polycystin-1, respectively. Single-channel studies of PC-2, S812A, and a substitution mutant, T721A, not related to phosphorylation show that PC-2 and S812A function as divalent cation channels with similar current amplitudes across a range of holding potentials; the T721A channel is not functional. Channel open probabilities for PC-2 and S812A show a bell-shaped dependence on cytoplasmic Ca 2؉ but there is a shift in this Ca 2؉ dependence such that S812A is 10-fold less sensitive to Ca 2؉ activation/inactivation than the wild type PC-2 channel. In vivo analysis of PC-2-dependent enhanced intracellular Ca 2؉ transients found that S812A resulted in enhanced transient duration and relative amplitude intermediate between control cells and those overexpressing wild type PC-2. Phosphorylation at Ser 812 modulates PC-2 channel activity and factors regulating this phosphorylation are likely to play a role in the pathogenesis of polycystic kidney disease.

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Essential Ca ²⁺ -Binding Motif for Ca ²⁺ -Sensitive Inactivation of L-Type Ca ²⁺ Channels

Cited by 261 publications

References 36 publications

Calcium-dependent inactivation of neuronal calcium channel currents is independent of calcineurin

Calcium-dependent inactivation of neuronal calcium channel currents is independent of calcineurin

Specific modulation of calmodulin activity induces a dramatic production of superoxide by alveolar macrophages

Calcium Dependence of Polycystin-2 Channel Activity Is Modulated by Phosphorylation at Ser812

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Essential Ca 2+ -Binding Motif for Ca 2+ -Sensitive Inactivation of L-Type Ca 2+ Channels

Cited by 261 publications

References 36 publications

Calcium-dependent inactivation of neuronal calcium channel currents is independent of calcineurin

Calcium-dependent inactivation of neuronal calcium channel currents is independent of calcineurin

Specific modulation of calmodulin activity induces a dramatic production of superoxide by alveolar macrophages

Calcium Dependence of Polycystin-2 Channel Activity Is Modulated by Phosphorylation at Ser812

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Essential Ca ²⁺ -Binding Motif for Ca ²⁺ -Sensitive Inactivation of L-Type Ca ²⁺ Channels