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FlaG binds KCNA5. 155 / 155
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"However, we found that bepridil inhibited the proteasome activity only at a very high concentration ( Fig. 6 ), suggesting that the stabilization of Kv1.5-FLAG by bepridil is independent of the protea[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23026372
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"We found that the EC 50 of bepridil-induced increases in Kv1.5-FLAG was about 1 μM, i.e., lower than the IC 50 of the acute blocking effect."
23026372
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"African green monkey kidney fibroblast (COS7) cells were used for transient expression of Kv1.5-FLAG and to study the effects of EPA and DHA on Kv1.5-FLAG protein stability."
19121632
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"Treatment with 4AP at 10–1000 μM for 24 h increased Kv1.5-FLAG proteins in a concentration-dependent manner ( Fig. 7 A)."
23026372
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"Fig. 7 B shows the levels of Kv1.5-FLAG in cells pretreated with 4AP alone or together with bepridil."
23026372
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"Whereas 4AP increased Kv1.5-FLAG proteins, an addition of 1 μM bepridil failed to further increase the Kv1.5-FLAG level in the presence of 100 μM 4AP."
23026372
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"Thus, the olprinone-induced stabilization of Kv1.5-FLAG proteins and enhancement of Kv1.5-FLAG currents might be due to phosphorylation of Kv1.5 proteins by activated PKA."
26368666
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"However, 8-bromo-cAMP failed to stabilize Kv1.5-FLAG proteins."
26368666
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"The Kv1.5 channel blocker 4AP, which induces conformational changes of Kv1.5 proteins and enhances their expression, abolished the increase of Kv1.5-FLAG protein expression by olprinone."
26368666
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"We found that the chronic treatment with olprinone at 30–1000 nM could stabilize Kv1.5-FLAG proteins."
26368666
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"Anti-FLAG staining of transfected COS cells revealed localization of Kv1.5-FLAG in a reticular pattern throughout the cytosol, obviously in the endoplasmic reticulum (ER)."
16185660
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"Kv1.5-FLAG was also localized in the Golgi apparatus as shown by its co-localization with Golgi-EYFP ( Fig. 3 A)."
16185660
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"We found no co-localization of Kv1.5-FLAG and Endosome-EGFP (data not shown)."
16185660
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"The interaction between Kv1.5-FLAG and CHIP was further demonstrated by 72.7% colocalization in the immunofluorescence experiment (supplementary Fig. 1)."
26232501
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"MG132 treatment (50 μM for 12 h) apparently increased the Kv1.5-FLAG immunoreactivity both in the ER and Golgi apparatus ( Fig. 3 A)."
16185660
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"To examine whether MG132 also altered the levels of cell-surface Kv1.5-FLAG, we examined Ik ur currents."
16185660
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"Depolarizing test pulses elicited outward currents in cells expressing Kv1.5-FLAG and these currents were partially blocked by 4-AP."
16185660
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"Regardless of MG132 treatment, we found no 4-AP-sensitive currents in cells transfected with an empty vector plasmid (data not shown), indicating that the observed Ik ur currents were totally derived [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
16185660
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"The kinetics of Ik ur currents through Kv1.5-FLAG were indistinguishable from those through Kv1.5, indicating no interference of the FLAG tag with the channel activity (data not shown)."
16185660
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"Since our immunofluorescence experiments showed a significant increase in Kv1.5-FLAG proteins in the ER and Golgi apparatus of the cells treated with MG132, it was likely that the MG132-induced increa[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
16185660
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"African green monkey kidney fibroblast (COS7) cells were used for transient expression of Kv1.5-FLAG."
23026372
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"The main site of this action appeared to be the ER: olprinone increased the signals of Kv1.5-FLAG proteins in the ER and Golgi apparatus, suggesting that the enhancement of Kv1.5-FLAG expression by ol[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"They were transfected with Kv1.5-FLAG cDNA together with enhanced green fluorescent protein (EGFP) constructs (pEGFP) using lipofectamine (Gibco BRL), and 48 h after transfection, cells were subjected[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23026372
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"To analyze the degradation kinetics of Kv1.5-FLAG, COS7 cells were seeded in 6-well plates and transfected with the Kv1.5-FLAG plasmid."
23026372
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"The distributions of Kv1.5-FLAG, AcGFP-ER, AcGFP-Golgi, and AcGFP-Mem were examined using Zeiss LSM700 (Deutschland, Germany)."
23026372
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"We first examined the effects of bepridil at various concentrations (0.1–30 μM) on the protein level of Kv1.5-FLAG expressed in COS7 cells."
23026372
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"Western blot analysis showed that bepridil increased Kv1.5-FLAG protein levels in a concentration-dependent manner ( Fig. 1 A)."
23026372
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"The increase in protein levels by bepridil was not due to bepridil-induced changes in the protein solubility, because Kv1.5-FLAG was present only in the detergent-soluble fraction of both control and [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23026372
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"Degradation of Kv1.5-FLAG was significantly delayed by both bepridil and MG132."
23026372
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"When expressed in COS cells, Kv1.5-FLAG had a relatively short half-life time and a part of the protein was ubiquitinated."
16185660
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"Lysosomal inhibitor, chloroquine (400 nM), did not influence the stability of Kv1.5-FLAG (Supplementary material Fig. S2 )."
23026372
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"This stabilization of Kv1.5-FLAG by proteasomal inhibition has been observed in the primary cultured atrial myocytes ( Fig. 2 )."
16185660
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"Next, we examined the intracellular localization of Kv1.5-FLAG expressed in COS7 cells in the absence and presence of bepridil."
23026372
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"Together with the localization of expressed proteins in the ER/Golgi apparatus but not in the endosome ( Fig. 3 ), these results strongly argue that Kv1.5-FLAG was degraded in the cytosol by the prote[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
16185660
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"Kv1.5-FLAG localized in the ER and Golgi apparatus, as shown by its colocalization with AcGFP-ER and AcGFP-Golgi, respectively."
23026372
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"Signals from markers co-localized with Kv1.5-FLAG signals up to 98.2% (Supplementary material Fig. S3 )."
23026372
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"Bepridil at 3 μM increased the signals of Kv1.5-FLAG both in the ER and Golgi apparatus ( Fig. 3 A and B)."
23026372
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"Fig. 3 C shows the localization of Kv1.5-FLAG and the cell surface marker AcGFP-Mem."
23026372
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"The signals of Kv1.5-FLAG superimposed on AcGFP-Mem were much higher in the presence of 3 μM bepridil than in its absence."
23026372
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"Fig. 3 D shows the results of quantitative analysis of the density of Kv1.5-FLAG in the ER, Golgi apparatus, and cell membrane."
23026372
sparser
"Treatment with bepridil significantly increased the signals of Kv1.5-FLAG in the ER, Golgi apparatus, and cell membrane, suggesting that the main site of bepridil action is the ER."
23026372
sparser
"Because it prolonged the half-life of the Kv1.5 protein, and its action was mimicked by a proteasome inhibitor ( Fig. 2 ), it appeared to stabilize Kv1.5-FLAG proteins by inhibiting their degradation."
23026372
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"The main site of its action appeared to be the ER, because immunofluorescence study showed that bepridil increased the density of Kv1.5-FLAG in the ER, Golgi apparatus, and the cell membrane ( Figs."
23026372
sparser
"In accordance with the result from the immunofluorescence study, cell fraction experiments showed that bepridil increased the Kv1.5-FLAG protein in both ER fractions and membrane fractions ( Fig. 4 )."
23026372
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"To examine whether bepridil increases the density of functional Kv1.5-FLAG on the cell surface, we measured the Kv1.5-mediated membrane current."
23026372
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"In cells expressing Kv1.5-FLAG, depolarizing test pulses elicited outward currents, which were almost completely blocked by 4AP at 1 mM."
23026372
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"Regardless of a treatment with bepridil, we found no 4AP-sensitive currents in cells transfected with an empty vector plasmid (data not shown), indicating that the observed currents were totally deriv[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
23026372
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"Kinetics of the current through Kv1.5-FLAG were nearly identical to that of Kv1.5, indicating no interference of the FLAG tag with the channel activity (Supplementary material Fig. S6 )."
23026372
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"Based on our previous report ( Kato et al., 2005 ), the half life of the newly synthesized Kv1.5-FLAG was estimated to be 6 h, most of Kv1.5-FLAG being degraded within 12 h."
19121632
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"EPA at 10 μM significantly increased Kv1.5-FLAG, while EPA at 100 μM significantly decreased it."
19121632
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"In contrast, DHA at 10–100 μM significantly decreased Kv1.5-FLAG protein in a dose-dependent manner."
19121632
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"Fig. 3 shows the chronic effects of EPA and DHA at 0.1–10 μM on Kv1.5-FLAG expression in COS7 cells."
19121632
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"EPA at the lower concentrations significantly increased Kv1.5-FLAG in a dose-dependent manner, indicating that EPA has chronic enhancing effects on Kv1.5 expression at lower concentrations."
19121632
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"In contrast, DHA at the same concentrations as EPA did not increase Kv1.5-FLAG."
19121632
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"We further examined the chronic effects of EPA on the stability of Kv1.5-FLAG using a degradation assay."
19121632
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"Fig. 5 shows the degradation process of Kv1.5-FLAG after treatment with cycloheximide (60 μg/ml) in the presence or absence of 1 μM EPA."
19121632
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"The half-time ( t 1/2 ) of Kv1.5-FLAG was 6.7 ± 1.8 h ( n = 4) in the control, and 14.1 ± 2.2 h ( n = 4: p < 0.05) in the presence of EPA, indicating that EPA stabilized Kv1.5-FLAG."
19121632
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"We then examined the intracellular localization of Kv1.5-FLAG expressed in COS7 cells in the absence and presence of 1 μM EPA."
19121632
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"Kv1.5-FLAG localized in the endoplasmic reticulum as shown by its co-localization with ER-EYFP ( Fig. 6 A , panels a–d)."
19121632
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"Kv1.5-FLAG also localized in the Golgi apparatus as shown by its co-localization with Golgi-EYFP ( Fig. 6 A, panels e–h)."
19121632
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"We found no co-localization of Kv1.5-FLAG and endosome-EGFP, in accordance with the lacking effect of the lysosome inhibitor chloroquine (2 mM) on the stability of Kv1.5-FLAG (data not shown)."
19121632
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"EPA at 1 μM remarkably increased the signal of Kv1.5-FLAG in the endoplasmic reticulum ( Fig. 6 A, panels b and d) and Golgi apparatus ( Fig. 6 A, panels f and h)."
19121632
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"Fig. 6 B shows the localization of Kv1.5-FLAG (panels a and d) and the cell surface marker AcGFP-Mem (panels b and e) in the transfected cells."
19121632
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"The signal of Kv1.5-FLAG on the cell surface to be superimposed on AcGFP-Mem was much larger in the presence of 1 μM EPA (panel f) than in the absence of EPA (panel c)."
19121632
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"Fig. 7 A shows the density of Kv1.5-FLAG in the endoplasmic reticulum (panel a), Golgi apparatus (panel b) and cell surface (panel c) determined using the individual marker GFP in the cells."
19121632
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"Treatment with 1 μM EPA significantly increased the distribution of Kv1.5-FLAG in the endoplasmic reticulum, Golgi apparatus and cell membrane, suggesting that the main sites of the stabilization of K[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
19121632
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"As shown in Fig. 7 B, treatment with EPA at 10 μM increased the Kv1.5-FLAG protein in both the endoplasmic reticulum fractions (#11 to 13) and the membrane fractions (#2 to 5), while Kv1.5-FLAG protei[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
19121632
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"Fig. 8 B shows the expression of Kv1.5-FLAG in the cells pretreated with the Kv1.5 current blocker 4AP at 1 mM alone or with both EPA (10 μM) and 4AP."
19121632
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"Although 4AP increased Kv1.5-FLAG protein significantly, addition of EPA (10 μM) did not influence the 4AP-induced increment of Kv1.5-FLAG."
19121632
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"In the cells co-transfected with both Kv1.5-FLAG and SAP97, EPA at 1 and 10 μM increased but EPA at 30 and 300 μM decreased the Kv1.5-FLAG expression ( Supplemental Fig. S4 C)."
19121632
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"From the aspect of transcript abundance of Kv1.5, we finally examined the effect of EPA on the mRNA of Kv1.5-FLAG in transfected COS7 cells."
19121632
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"In the present study, chronic treatment with EPA at lower concentrations for 12 h was sufficient for a post-translational modification of Kv1.5-FLAG and its stabilization."
19121632
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"In immunocytochemical data, EPA increased the density of Kv1.5-FLAG in the endoplasmic reticulum and Golgi apparatus, suggesting that the increase of channel activity by EPA was most likely secondary [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
19121632
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"This finding was confirmed by the EPA-induced increase in the distribution of Kv1.5-FLAG in the endoplasmic reticulum and membrane fractions ( Fig. 7 B)."
19121632
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"The molecular basis underlying the EPA-induced post-translational modification of Kv1.5-FLAG and the difference from that induced by DHA remains to be determined."
19121632
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"In addition, EPA at lower concentrations increased endogenous and transfected SAP97 expression ( Fig. S4 ), suggesting the EPA stabilization of Kv1.5-FLAG via the increase of SAP97."
19121632
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"To study the effects of Hsps on Kv1.5-FLAG or HERG-FLAG protein stability, we used COS7 cells or HEK293 cells for transient expression and HeLa cells stably expressing heat shock factor 1 (HeLa/HSF-1)[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
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"COS7 cells or HEK293 cells were co-transfected with Kv1.5-FLAG cDNA (2 μg) or HERG-FLAG cDNA (2 μg) and either pcDNA3/Hsp70 (2 μg) or pcDNA3/Hsp90 (2 μg) together with pEGFP (0.5 μg), keeping the tota[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
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"The surface plot module of National Institute of Health Image 1.01 free ware was used for integrated density volume mapping to quantitatively assess the intracellular abundance of Kv1.5-FLAG, ER-EYFP,[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
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"Kv1.5-FLAG was transiently expressed in HeLa cells stably expressing HSF-1, Hsp27, Hsp70, and Hsp40, and was detected by Western blotting ( Fig. 1 A)."
18502196
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"The protein level of Kv1.5-FLAG was higher in cells stably expressing HSF-1 or Hsp70 than in cells expressing Hsp27 or Hsp40."
18502196
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"Densitometry showed a significant increase in the ratio of Kv1.5-FLAG to GFP in cells over-expressing HSF-1 or Hsp70 ( Supplemental Fig. S1 )."
18502196
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"In contrast, transient expression of Hsp90 did not increase the level of Kv1.5-FLAG ( Fig. 1 C), and inhibition of Hsp90 function by geludanamycin failed to affect the level of Kv1.5-FLAG (data not sh[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
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"These results suggested that either over-expression of Hsp70 or induction of this molecular chaperone by HSF-1 stabilized Kv1.5-FLAG."
18502196
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"As shown in Fig. 2 A, Hsp70 increased the level of Kv1.5-FLAG."
18502196
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"Kv1.5-FLAG was recovered in the detergent-soluble fraction regardless of co-expression of Hsp70, excluding Hsp70-induced changes in protein solubility ( Fig. 2 B)."
18502196
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"Abundance of mRNA of Kv1.5-FLAG was comparable between cells co-transfected with and without Hsp70 ( Supplemental Fig. S2 )."
18502196
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"Immunoprecipitation showed that Hsp70 did not change the level of ubiquitinated Kv1.5-FLAG, while MG132 markedly increased the level of ubiquitinated Kv1.5-FLAG (data not shown)."
18502196
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"The t 1/2 of Kv1.5-FLAG was 6.7 ± 1.8 h ( n = 4) in control, and was prolonged to 14.1 ± 2.2 h ( n = 4) when co-transfected with Hsp70 ( Fig. 2 C)."
18502196
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"We next examined the interaction of Kv1.5-FLAG and Hsp70 in cells transfected with both plasmids."
18502196
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"Hsp70 was co-precipitated with Kv1.5-FLAG ( Supplemental Fig. S3A )."
18502196
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"We examined the distribution of Kv1.5-FLAG and Hsp70 in the transfected COS7 cells by immunofluorescence."
18502196
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"Anti-FLAG staining revealed the localization of Kv1.5-FLAG in a reticular pattern throughout the cytosol and in a perinuclear pattern throughout the nuclear envelope, indicating its presence in the en[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
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"In cells transfected with Hsp70, the signals of both Kv1.5-FLAG and Hsp70 remarkably increased ( Supplemental Fig. S5-d and e ) and co-localized with each other ( Supplemental Fig. S5-f )."
18502196
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"We then compared intracellular localization of Kv1.5-FLAG in COS7 cells transfected with or without Hsp70."
18502196
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"Kv1.5-FLAG was localized in the Golgi apparatus as shown by its co-localization with Golgi-EYFP ( Supplemental Fig. S6-a and b )."
18502196
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"We found no co-localization of Kv1.5-FLAG and Endosome-EGFP (data not shown), in accordance with the lack of effects of a lysosome inhibitor chloroquine (2 mM) on the stability of Kv1.5-FLAG (data not[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
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"Hsp70 over-expression remarkably increased the signal of Kv1.5-FLAG in the Golgi apparatus ( Supplemental Fig. S6-c and d ) and ER ( Supplemental Fig. S6-g and h )."
18502196
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"Fig. 3 shows the localization of Kv1.5-FLAG (panels A and D) and AcGFP-Mem, a cell surface marker (panels B and E)."
18502196
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"While the signal of Kv1.5-FLAG did not co-localize with AcGFP-Mem (panel C) in cells expressing endogenous Hsp70, Kv1.5-FLAG co-localized with it (panel F) in cells over-expressing Hsp70."
18502196
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"We performed a quantitative analysis of Kv1.5-FLAG in the ER, Golgi apparatus and on the cell surface ( Supplemental Fig. S7 )."
18502196
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"Hsp70 over-expression significantly increased the signals of Kv1.5-FLAG in all of these compartments."
18502196
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"In cells transfected with Kv1.5-FLAG alone, Kv1.5-FLAG and an ER marker, calnexin, were predominantly recovered in fractions 10–16, while a membrane marker Na + /K + -ATPase was enriched in fractions [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
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"In cells transfected with both Kv1.5-FLAG and Hsp70, Kv1.5-FLAG was broadly located in fraction 2–16 ( Supplemental Fig. S8 )."
18502196
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"To examine whether Hsp70 altered the density of functional Kv1.5-FLAG on the cell surface, we measured the Kv1.5-mediated membrane current."
18502196
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"In cells expressing Kv1.5-FLAG, depolarizing test pulses elicited outward currents, which were almost completely blocked by 4-AP."
18502196
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"Regardless of co-expression of Hsp70, we found no 4-AP-sensitive currents in cells transfected with an empty vector plasmid (data not shown), indicating that the observed current was totally derived f[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
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"Kinetics of the current through Kv1.5-FLAG were nearly identical to that through Kv1.5, indicating no interference of the FLAG tag with the channel activity (data not shown)."
18502196
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"Since our immunofluorescence experiments showed a significant increase in Kv1.5-FLAG protein in the Golgi apparatus, ER and on the cell surface of cells over-expressing Hsp70, it was likely that the H[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
sparser
"In the present study, we found that over-expression of Hsp70 increased the level of Kv1.5-FLAG protein ( Figs."
18502196
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"Both co-immunoprecipitation ( Supplemental Fig. S3A ) and co-localization ( Fig. 3 , Supplemental Fig. S6 ) experiments suggested that Hsp70 could interact with Kv1.5-FLAG."
18502196
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"Other Hsp proteins, Hsp27, Hsp40, and Hsp90, failed to affect the level of Kv1.5-FLAG ( Fig. 1 , Supplemental Fig. S1 ), indicating that the stabilizing effect was unique to Hsp70."
18502196
sparser
"In the present study, over-expression of Hsp90 increased the mature form of HERG ( Fig. 1 B) but did not affect the level of Kv1.5-FLAG ( Fig. 1 C)."
18502196
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"The molecular basis underlying the interaction between Hsp70 and Kv1.5-FLAG remains to be determined."
18502196
sparser
"In the present study, over-expression of Hsp70 increased the amount of Kv1.5-FLAG in the ER and Golgi apparatus ( Supplemental Fig. S6 ) as well as on the cell surface ( Fig. 3 ), which was consistent[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
18502196
sparser
"African green monkey kidney fibroblast (COS7) cells were used for transient expression of Kv1.5-FLAG and to study effects of olprinone on Kv1.5-FLAG protein stability."
26368666
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"Kv1.5-FLAG expressed in COS cells was detected by anti-FLAG Western blotting at ∼80 kDa, a predicted size of full-length Kv1.5-FLAG."
16185660
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"To analyze the degradation kinetics of Kv1.5-FLAG, COS7 cells were seeded in 6-well plates and transfected with pRC/Kv1.5-FLAG."
26368666
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"Treatment with MG132 (50 μM for 12 h) increased the density of this band ( Fig. 1 A), suggesting MG132-induced stabilization of Kv1.5-FLAG."
16185660
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"The distributions of Kv1.5-FLAG, AcGFP-ER, AcGFP-Golgi, AcGFP-Mem and endosome-EGFP were examined using a Zeiss LSM700 confocal microscope illuminated with a krypton/argon laser (Deutschland, Germany)[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"Kv1.5-FLAG was recovered in the detergent-soluble fraction regardless of MG132 treatment, excluding drug-induced changes in protein solubility ( Fig. 1 B)."
16185660
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"Immunoprecipitation experiments showed that MG132 treatment increased levels of ubiquitinated Kv1.5-FLAG ( Fig. 1 D)."
16185660
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"Effects of olprinone on protein levels of Kv1.5-FLAG were examined in transfected COS7 cells."
26368666
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"Fig. 1 A shows a representative western blot of Kv1.5-FLAG proteins."
26368666
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"Olprinone increased the level of Kv1.5-FLAG proteins in a concentration-dependent manner with the maximum effect attained at 300 nM, as summarized in Fig. 1 B. Effects of olprinone on the stability of[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"Olprinone at 100 nM significantly slowed the degradation of Kv1.5-FLAG proteins ( Fig. 2 ): the t 1/2 values in the absence and presence of olprinone was 6.6±1.0 h ( n =4) and 11.8±1.5 h ( n =4; P <0.[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"Intracellular localization of Kv1.5-FLAG proteins was examined by immuno- fluorescence."
26368666
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"To confirm MG132-induced stabilization of Kv1.5-FLAG, we determined a half-life time ( t 1/2 ) of expressed proteins by pulse-chase analyses."
16185660
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"Signals of Kv1.5-FLAG proteins were detected on the plasma membrane, ER and Golgi apparatus as shown in Fig. 3 A by their co-localization with AcGFP-Mem (panels a–c), AcGFP-ER (panels d–f), and AcGFP-[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"In cells expressing Kv1.5-FLAG, depolarizing test pulses elicited outward currents, which were almost completely blocked by 1 mM 4AP."
26368666
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"Treatment with olprinone at 100 nM for 12 h increased the amplitude of expressed Kv1.5-FLAG currents ( Fig. 4 A)."
26368666
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"Kv1.5-FLAG had a t 1/2 of 6.7 ± 1.2 h ( n = 4), and this value was prolonged to 20.2 ± 3.4 h ( n = 4) in cells treated with MG132 ( Fig. 1 E)."
16185660
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"As summarized in Fig. 4 B, olprinone at 100 nM significantly augmented Kv1.5-FLAG currents at the membrane potentials ranging from +20 to +80 mV without affecting the threshold potential of the curren[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"Fig. 4 C shows the representative original traces of tail currents recorded for determination of the steady-state voltage-dependent activation curve for Kv1.5-FLAG currents in transfected cells in the[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"Thus, olprinone increased the density of Kv1.5-FLAG currents without changing the voltage-dependent activation kinetics."
26368666
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"We further investigated effects of a protein transport inhibitor, colchicine s , on Kv1.5-FLAG currents."
26368666
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"Fig. 5 A shows representative traces of Kv1.5-FLAG currents recorded from cells treated with or without colchicine."
26368666
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"Treatment with this agent at 5 μM for 12 h significantly decreased the basal level of Kv1.5-FLAG currents and minimized the olprinone-induced increase of the currents as summarized in Fig. 5 B. Colchi[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"Fig. 6 A shows the levels of Kv1.5-FLAG proteins in cells pretreated with 4AP alone or together with olprinone."
26368666
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"4AP at 100 μM significantly increased the level of Kv1.5-FLAG proteins, indicating its chemical chaperone action."
26368666
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"Olprinone at 100 nM failed to further increase the Kv1.5-FLAG level in the presence of 100 μM 4AP."
26368666
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"As summarized in Fig. 6 B, 4AP abolished the enhancement of Kv1.5-FLAG protein expression by olprinone."
26368666
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"Thus, the stabilizing effect of olprinone on Kv1.5-FLAG channel proteins was inhibited by the chemical chaperone that directly binds to the Kv1.5 protein."
26368666
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"To determine whether an increase in intracellular cAMP could influence the protein level of Kv1.5 or not, we further examined effects of 8-bromo-cAMP on Kv1.5-FLAG protein levels."
26368666
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"8-bromo-cAMP at 20 μM did not significantly increase the protein level of Kv1.5-FLAG, nor did it inhibit the increasing effect of 100 nM olprinone on the Kv1.5 protein level ( Fig. 7 )."
26368666
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"Thus, the stabilizing effect of olprinone on Kv1.5-FLAG channel proteins was not attributable to its action of elevating intracellular cAMP."
26368666
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"In the present study, we found that (1) olprinone slowed the degradation of the Kv1.5-FLAG protein and increased its level in a concentration-dependent manner; (2) olprinone enhanced the signals of Kv[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"In the present study, olprinone increased immunofluorescent signals of Kv1.5-FLAG proteins on the plasma membrane and Kv1.5 currents without changes in their activation kinetics, which is a possible m[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"Next, we examined intracellular localization of Kv1.5-FLAG by immunofluorescence."
16185660
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"This notion is consistent with the effect of colchicine, which probably inhibited intracellular transport of mature Kv1.5-FLAG proteins from the ER/Golgi apparatus to the cell surface and abolished th[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
26368666
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"In brief, cells were maintained in Dulbecco's modified Eagle medium (Gibco BRL)/10% fetal bovine serum (Gibco BRL) at 37 °C in a 5% CO 2 incubator, and transfected with Kv1.5-FLAG cDNA (2 μg) together[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
19121632
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"To analyze the degradation kinetics of Kv1.5-FLAG, COS7 cells were seeded in 6-well plates and transfected with the Kv1.5-FLAG plasmid."
19121632
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"The distribution of Kv1.5-FLAG, ER-EYFP, Golgi-EYFP, AcGFP-Mem and endosome-EGFP was examined using a Bio-Rad MRC 1024 confocal microscope illuminated with a krypton/argon laser."
19121632
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"The surface plot module 1.01 Image freeware from the National Institutes of Health was used for integrated density volume mapping to quantify intracellular Kv1.5-FLAG, ER-EYFP, Golgi-EYFP, AcGFP-Mem a[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"
19121632
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"The chronic effects of EPA or DHA on Kv1.5-FLAG were examined in transfected COS7 cells ( Fig. 2 )."
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