Amino acid substitutions in the rat Na+, K(+)‐ATPase alpha 2‐subunit alter the cation regulation of pump current expressed in HeLa cells.

Yamamoto, Satoshi; Kuntzweiler, Theresa A.; Wallick, Earl T.; Sperelakis, Nicholas; Yatani, Atsuko

doi:10.1113/jphysiol.1996.sp021629

Cited by 21 publications

(14 citation statements)

References 27 publications

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“…Electrophysiological measurements on HeLa cells expressing the ␣2 mutant D925L have demonstrated that the electrogenicity of the pump is retained with this mutation; hence, the 3Na ϩ / 2K ϩ stoichiometry is probably also intact (56). It is noteworthy in this connection that the D923N mutant supported cell viability in our experiments, thus being able to transport Na ϩ and K ϩ and probably to generate a membrane potential.…”

Section: Discussionsupporting

confidence: 55%

The Rapid-onset Dystonia Parkinsonism Mutation D923N of the Na+,K+-ATPase α3 Isoform Disrupts Na+ Interaction at the Third Na+ Site

Einholm¹,

Toustrup-Jensen²,

Holm

et al. 2010

Journal of Biological Chemistry

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Rapid-onset dystonia parkinsonism (RDP), a rare neurological disorder, is caused by mutation of the neuron-specific ␣3-isoform of Na ؉ ,K ؉ -ATPase. Here, we present the functional consequences of RDP mutation D923N. Relative to the wild type, the mutant exhibits a remarkable ϳ200-fold reduction of Na ؉ affinity for activation of phosphorylation from ATP, reflecting a defective interaction of the E 1 form with intracellular Na ؉ . This is the largest effect on Na ؉ affinity reported so far for any Na ؉ ,K ؉ -ATPase mutant. D923N also affects the interaction with extracellular Na ؉ normally driving the E 1 P to E 2 P conformational transition backward. However, no impairment of K ؉ binding was observed for D923N, leading to the conclusion that Asp 923 is specifically associated with the third Na ؉ site that is selective toward Na ؉ . The crystal structure of the Na ؉ ,K ؉ -ATPase in E 2 form shows that Asp 923 is located in the cytoplasmic half of transmembrane helix M8 inside a putative transport channel, which is lined by residues from the transmembrane helices M5, M7, M8, and M10 and capped by the C terminus, recently found involved in recognition of the third Na ؉ ion. Structural modeling of the E 1 form of Na ؉ ,K ؉ -ATPase based on the Ca 2؉ -ATPase crystal structure is consistent with the hypothesis that Asp 923 contributes to a site binding the third Na ؉ ion. These results in conjunction with our previous findings with other RDP mutants suggest that a selective defect in the handling of Na ؉ may be a general feature of the RDP disorder.

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Section: Discussionsupporting

confidence: 55%

The Rapid-onset Dystonia Parkinsonism Mutation D923N of the Na+,K+-ATPase α3 Isoform Disrupts Na+ Interaction at the Third Na+ Site

Einholm¹,

Toustrup-Jensen²,

Holm

et al. 2010

Journal of Biological Chemistry

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“…Whole-cell patch clamp studies were performed as described previously (17,(21)(22)(23). Cell capacitance was measured using voltage ramps of 0.8V/s from a holding potential of −50 mV.…”

Section: Electrophysiological Recordingsmentioning

confidence: 99%

“…All experiments were performed at room temperature (20°C-22°C). The experimental chamber (0.2 mL) was placed on a microscope stage, and external solution changes were made rapidly using a modified Y-tube technique (21). The Na + /Ca 2+ exchange currents were recorded as previously described.…”

Section: Electrophysiological Recordingsmentioning

confidence: 99%

Mesenteric Lymph From Rats With Thermal Injury Prolongs the Action Potential and Increases Ca2+ Transient in Rat Ventricular Myocytes

Yatani¹,

Xu²,

Kim³

et al. 2003

Shock

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Although gut-derived mesenteric lymph from animals with thermal injury appears to lead to myocardial contractile dysfunction, the cellular mechanisms remain unclear. We examined the direct effects of intestinal lymph on excitation-contraction coupling in rat ventricular myocytes. Lymph from rats receiving burn injury (burn lymph), but not from sham-burned rats, rapidly enhanced myocyte contraction and the amplitude of Ca2+ transient; the average percentage of shortening was increased from 5.5 +/- 0.3% to 10.5 +/- 0.9%. 90% and the Ca2+ transients increased by 80% +/- 20%. Burn lymph had no effect on the amplitude of L-type Ca2+ current (ICa) or the inward rectifier K+ current, but the transient outward K+ currents (Ito) were reduced significantly by burn lymph. Inhibition of Ito was not altered by an alpha1-adrenergic receptor (AR) antagonist, prazosin, indicating that the block was not mediated via alpha1-AR signaling pathway. Action potential (AP) duration, measured at 50% and 90% repolarization, was prolonged by burn lymph. Stimulation of myocytes with AP voltage-clamp waveforms derived from prolonged AP induced by burn lymph revealed a 1.7-fold increase in Ca2+ influx via ICa compared with the Ca2+ influx induced by control AP. Blocking of Ito by 4-aminopyridine prolonged AP duration and increased Ca2+ transients, mimicking the effects of burn lymph. Burn lymph did not affect Na+/Ca2+ exchange currents or caffeine-induced SR Ca2+ release. Thus, acute exposure of normal cardiac myocytes to burn lymph increases Ca2+ transients by a prolongation of AP as a result of a reduction of Ito with no intrinsic change in ICa or exchanger. The electrophysiological changes are similar to those that occur during compensated cardiac hypertrophy, suggesting a common mechanistic link between burn lymph- and hypertrophy-induced cardiac dysfunction.

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“…The patch pipettes had a resistance of 2 MΩ or less. The experimental chamber (0.2 ml) was placed on a microscope stage, and the external solution changes were made using a modified Y-tube technique (24 3). These external and internal solutions provided isolation of Ca 2+ channel currents (I Ca ) from other membrane currents, such as Na + and K + channel currents, and also from Ca 2+ flux through the Na + /Ca 2+ exchanger (25).…”

Section: Figurementioning

confidence: 99%

Differential regulation of inotropy and lusitropy in overexpressed Gsα myocytes through cAMP and Ca2+ channel pathways

Kim

Yatani²,

Vatner³

et al. 1999

J. Clin. Invest.

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We investigated the mechanisms responsible for altered contractile and relaxation function in overexpressed Gsα myocytes. Although baseline contractile function (percent contraction) in Gsα mice was similar to that of wild-type (WT) mice, left ventricular myocyte contraction, fura-2 Ca 2+ transients, and Ca 2+ channel currents (I ca ) were greater in Gsα mice in response to 10 -8 M isoproterenol (ISO) compared with WT mice. The late phase of relaxation of the isolated myocytes and fura-2 Ca 2+ transients was accelerated at baseline in Gsα but did not increase further with ISO. In vivo measurements using echocardiography also demonstrated enhanced relaxation at baseline in Gsα mice. Forskolin and CaCl 2 increased contraction similarly in WT and Gsα mice. Rp-cAMP, an inhibitor of protein kinase, blocked the increases in contractile response and Ca 2+ currents to ISO in WT and to forskolin in both WT and Gsα. It also blocked the accelerated relaxation in Gsα at baseline but not the contractile response to ISO in Gsα myocytes. Baseline measurements of cAMP and phospholambation phosphorylation were enhanced in Gsα compared with WT. These data indicate that overexpression of Gsα accelerates relaxation at end diastolic but does not affect baseline systolic function in isolated myocytes. However, the enhanced responses to sympathetic stimulation partly reflect increased Ca 2+ channel activity; i.e the cellular mechanisms mediating these effects appear to involve a cAMP-independent as well as a cAMPdependent pathway.

show abstract

Amino acid substitutions in the rat Na+, K(+)‐ATPase alpha 2‐subunit alter the cation regulation of pump current expressed in HeLa cells.

Cited by 21 publications

References 27 publications

The Rapid-onset Dystonia Parkinsonism Mutation D923N of the Na+,K+-ATPase α3 Isoform Disrupts Na+ Interaction at the Third Na+ Site

The Rapid-onset Dystonia Parkinsonism Mutation D923N of the Na+,K+-ATPase α3 Isoform Disrupts Na+ Interaction at the Third Na+ Site

Mesenteric Lymph From Rats With Thermal Injury Prolongs the Action Potential and Increases Ca2+ Transient in Rat Ventricular Myocytes

Differential regulation of inotropy and lusitropy in overexpressed Gsα myocytes through cAMP and Ca2+ channel pathways

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