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Cellular mechanisms of mutations in Kv7.1: auditory functions in Jervell and Lange-Nielsen syndrome vs. Romano-Ward syndrome.

Mousavi Nik A, Gharaie S, Jeong Kim H - Front Cell Neurosci (2015)

Bottom Line: As a result of cell-specific functions of voltage-activated K(+) channels, such as Kv7.1, mutations in this channel produce profound cardiac and auditory defects.The RWS mutants exhibited varied functional phenotypes.However, they can be summed up as exhibiting DN effects.

View Article: PubMed Central - PubMed

Affiliation: Department of Anesthesiology and Pain Medicine, Center for Neuroscience, School of Medicine, University of California, Davis Davis, CA, USA.

ABSTRACT
As a result of cell-specific functions of voltage-activated K(+) channels, such as Kv7.1, mutations in this channel produce profound cardiac and auditory defects. At the same time, the massive diversity of K(+) channels allows for compensatory substitution of mutant channels by other functional channels of their type to minimize defective phenotypes. Kv7.1 represents a clear example of such functional dichotomy. While several point mutations in the channel result in a cardio-auditory syndrome called Jervell and Lange-Nielsen syndrome (JLNS), about 100-fold mutations result in long QT syndrome (LQTS) denoted as Romano-Ward syndrome (RWS), which has an intact auditory phenotype. To determine whether the cellular mechanisms for the diverse phenotypic outcome of Kv7.1 mutations, are dependent on the tissue-specific function of the channel and/or specialized functions of the channel, we made series of point mutations in hKv7.1 ascribed to JLNS and RWS. For JLNS mutations, all except W248F yielded non-functional channels when expressed alone. Although W248F at the end of the S4 domain yielded a functional current, it underwent marked inactivation at positive voltages, rendering the channel non-functional. We demonstrate that by definition, none of the JLNS mutants operated in a dominant negative (DN) fashion. Instead, the JLNS mutants have impaired membrane trafficking, trapped in the endoplasmic reticulum (ER) and Cis-Golgi. The RWS mutants exhibited varied functional phenotypes. However, they can be summed up as exhibiting DN effects. Phenotypic differences between JLNS and RWS may stem from tissue-specific functional requirements of cardiac vs. inner ear non-sensory cells.

No MeSH data available.


Related in: MedlinePlus

Current phenotype of homomeric and heteromeric hKv7.1 channel and RWS mutants, R243P, L250H and G306V channels. (A) CHO cells were transfected with RWS mutant R243P DNA. Whole-cell outward currents were recorded after 48 h. Cells were held at −80 mV and stepped to voltages ranging from −100 to 60 mV using Δ V = 10 mV. Homomeric MT channel R243P did not yield outward currents (left panel). Similar currents were obtained for homomeric L250H and G306V MTs. However, co-expression WT hKv7.1 and MT R243P at a ratio of 2:2 (middle panel) yielded reduced current magnitude (middle panel), which was enhanced further after increasing the ratio of the WT-hKv7.1:R243P (3:1, right panel). (B) Plots of representative current density-voltage relation of currents derived from WT hKv7.1 alone () and a combination (ratio 2:2) of WT Kv7.1 and MTs (MT-R243P ◻, MT-L250H ◾, and MT-G306V ○). (C) Co-expression of WT hKv7.1 and MT-R243P, L250H and G306V at a ratio (2:2) produced a right-ward shift in the voltage-dependent activation of the resulting currents. Steady-state activation curves of hKv7.1 alone () and after suppression by co-expression with the MT channels are shown. Tail currents were measured at −40 mV, and normalized to the largest tail current magnitude, and plotted against the preceding pre-pulse voltages. The midpoint (V1/2, in mV) and the slope factors (k, in mV) were as follows: V1/2 and k for WT-hKv7.1 alone and in combination WT-hKv7.1:R243P, WT-hKv7.1:L250H, and WT-hKv7.1:G306V were −15.8 ± 1.2, 11.8 ± 0.9; 2.5 ± 1.1, 12.4 ± 0.6; 7.6 ± 2.2, 12.7 ± 1.0; 15.7 ± 2.4, 12.8 ± 1.6 mV (n = 11), respectively.
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Figure 7: Current phenotype of homomeric and heteromeric hKv7.1 channel and RWS mutants, R243P, L250H and G306V channels. (A) CHO cells were transfected with RWS mutant R243P DNA. Whole-cell outward currents were recorded after 48 h. Cells were held at −80 mV and stepped to voltages ranging from −100 to 60 mV using Δ V = 10 mV. Homomeric MT channel R243P did not yield outward currents (left panel). Similar currents were obtained for homomeric L250H and G306V MTs. However, co-expression WT hKv7.1 and MT R243P at a ratio of 2:2 (middle panel) yielded reduced current magnitude (middle panel), which was enhanced further after increasing the ratio of the WT-hKv7.1:R243P (3:1, right panel). (B) Plots of representative current density-voltage relation of currents derived from WT hKv7.1 alone () and a combination (ratio 2:2) of WT Kv7.1 and MTs (MT-R243P ◻, MT-L250H ◾, and MT-G306V ○). (C) Co-expression of WT hKv7.1 and MT-R243P, L250H and G306V at a ratio (2:2) produced a right-ward shift in the voltage-dependent activation of the resulting currents. Steady-state activation curves of hKv7.1 alone () and after suppression by co-expression with the MT channels are shown. Tail currents were measured at −40 mV, and normalized to the largest tail current magnitude, and plotted against the preceding pre-pulse voltages. The midpoint (V1/2, in mV) and the slope factors (k, in mV) were as follows: V1/2 and k for WT-hKv7.1 alone and in combination WT-hKv7.1:R243P, WT-hKv7.1:L250H, and WT-hKv7.1:G306V were −15.8 ± 1.2, 11.8 ± 0.9; 2.5 ± 1.1, 12.4 ± 0.6; 7.6 ± 2.2, 12.7 ± 1.0; 15.7 ± 2.4, 12.8 ± 1.6 mV (n = 11), respectively.

Mentions: Some KCNQ1 RWS mutants (R243P, L250H and G306V) were non-functional when transfected alone, but showed stunted currents when co-transfected with WT. The family of current traces in Figure 7A is an example of the phenotypic outcome of expression of R243P alone and at different ratios with the WT channel. The current density-voltage relationships of the three mutants suggest that although the mutant alone may be non-functional, the presence of the WT channel may render or facilitate mutant subunit functional (Figure 7B). The steady-state voltage-dependent activation of current generated from conditions when the WT:MT ratio is (2:2) were variable (Figure 7C). The midpoint (V1/2, in mV) and the slope factors (k, in mV) were as follows: V1/2 and k for WT-hKv7.1 alone and in combination (2:2) WT-hKv7.1:R243P, WT-Kv7.1:L250H, and WT-hKv7.1:G306V were −15.8 ± 1.2, 11.8 ± 0.9; 2.5 ± 1.1, 12.4 ± 0.6; 7.6 ± 2.2, 12.7 ± 1.0; 15.7 ± 2.4, 12.8 ± 1.6 mV (n = 11), respectively. The steady-state voltage-dependent activation was shifted rightward. Therefore, mutant channels were activated in more positive voltages in compare to WT (Figure 7C). Our results is in agreement with study conducted by Chouabe et al. (2000) where they showed that RWS mutants such as R243H and R533W not only reduced the amount of current, but also shifted the voltage-dependent activation of the channel to the more positive voltages.


Cellular mechanisms of mutations in Kv7.1: auditory functions in Jervell and Lange-Nielsen syndrome vs. Romano-Ward syndrome.

Mousavi Nik A, Gharaie S, Jeong Kim H - Front Cell Neurosci (2015)

Current phenotype of homomeric and heteromeric hKv7.1 channel and RWS mutants, R243P, L250H and G306V channels. (A) CHO cells were transfected with RWS mutant R243P DNA. Whole-cell outward currents were recorded after 48 h. Cells were held at −80 mV and stepped to voltages ranging from −100 to 60 mV using Δ V = 10 mV. Homomeric MT channel R243P did not yield outward currents (left panel). Similar currents were obtained for homomeric L250H and G306V MTs. However, co-expression WT hKv7.1 and MT R243P at a ratio of 2:2 (middle panel) yielded reduced current magnitude (middle panel), which was enhanced further after increasing the ratio of the WT-hKv7.1:R243P (3:1, right panel). (B) Plots of representative current density-voltage relation of currents derived from WT hKv7.1 alone () and a combination (ratio 2:2) of WT Kv7.1 and MTs (MT-R243P ◻, MT-L250H ◾, and MT-G306V ○). (C) Co-expression of WT hKv7.1 and MT-R243P, L250H and G306V at a ratio (2:2) produced a right-ward shift in the voltage-dependent activation of the resulting currents. Steady-state activation curves of hKv7.1 alone () and after suppression by co-expression with the MT channels are shown. Tail currents were measured at −40 mV, and normalized to the largest tail current magnitude, and plotted against the preceding pre-pulse voltages. The midpoint (V1/2, in mV) and the slope factors (k, in mV) were as follows: V1/2 and k for WT-hKv7.1 alone and in combination WT-hKv7.1:R243P, WT-hKv7.1:L250H, and WT-hKv7.1:G306V were −15.8 ± 1.2, 11.8 ± 0.9; 2.5 ± 1.1, 12.4 ± 0.6; 7.6 ± 2.2, 12.7 ± 1.0; 15.7 ± 2.4, 12.8 ± 1.6 mV (n = 11), respectively.
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Related In: Results  -  Collection

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Figure 7: Current phenotype of homomeric and heteromeric hKv7.1 channel and RWS mutants, R243P, L250H and G306V channels. (A) CHO cells were transfected with RWS mutant R243P DNA. Whole-cell outward currents were recorded after 48 h. Cells were held at −80 mV and stepped to voltages ranging from −100 to 60 mV using Δ V = 10 mV. Homomeric MT channel R243P did not yield outward currents (left panel). Similar currents were obtained for homomeric L250H and G306V MTs. However, co-expression WT hKv7.1 and MT R243P at a ratio of 2:2 (middle panel) yielded reduced current magnitude (middle panel), which was enhanced further after increasing the ratio of the WT-hKv7.1:R243P (3:1, right panel). (B) Plots of representative current density-voltage relation of currents derived from WT hKv7.1 alone () and a combination (ratio 2:2) of WT Kv7.1 and MTs (MT-R243P ◻, MT-L250H ◾, and MT-G306V ○). (C) Co-expression of WT hKv7.1 and MT-R243P, L250H and G306V at a ratio (2:2) produced a right-ward shift in the voltage-dependent activation of the resulting currents. Steady-state activation curves of hKv7.1 alone () and after suppression by co-expression with the MT channels are shown. Tail currents were measured at −40 mV, and normalized to the largest tail current magnitude, and plotted against the preceding pre-pulse voltages. The midpoint (V1/2, in mV) and the slope factors (k, in mV) were as follows: V1/2 and k for WT-hKv7.1 alone and in combination WT-hKv7.1:R243P, WT-hKv7.1:L250H, and WT-hKv7.1:G306V were −15.8 ± 1.2, 11.8 ± 0.9; 2.5 ± 1.1, 12.4 ± 0.6; 7.6 ± 2.2, 12.7 ± 1.0; 15.7 ± 2.4, 12.8 ± 1.6 mV (n = 11), respectively.
Mentions: Some KCNQ1 RWS mutants (R243P, L250H and G306V) were non-functional when transfected alone, but showed stunted currents when co-transfected with WT. The family of current traces in Figure 7A is an example of the phenotypic outcome of expression of R243P alone and at different ratios with the WT channel. The current density-voltage relationships of the three mutants suggest that although the mutant alone may be non-functional, the presence of the WT channel may render or facilitate mutant subunit functional (Figure 7B). The steady-state voltage-dependent activation of current generated from conditions when the WT:MT ratio is (2:2) were variable (Figure 7C). The midpoint (V1/2, in mV) and the slope factors (k, in mV) were as follows: V1/2 and k for WT-hKv7.1 alone and in combination (2:2) WT-hKv7.1:R243P, WT-Kv7.1:L250H, and WT-hKv7.1:G306V were −15.8 ± 1.2, 11.8 ± 0.9; 2.5 ± 1.1, 12.4 ± 0.6; 7.6 ± 2.2, 12.7 ± 1.0; 15.7 ± 2.4, 12.8 ± 1.6 mV (n = 11), respectively. The steady-state voltage-dependent activation was shifted rightward. Therefore, mutant channels were activated in more positive voltages in compare to WT (Figure 7C). Our results is in agreement with study conducted by Chouabe et al. (2000) where they showed that RWS mutants such as R243H and R533W not only reduced the amount of current, but also shifted the voltage-dependent activation of the channel to the more positive voltages.

Bottom Line: As a result of cell-specific functions of voltage-activated K(+) channels, such as Kv7.1, mutations in this channel produce profound cardiac and auditory defects.The RWS mutants exhibited varied functional phenotypes.However, they can be summed up as exhibiting DN effects.

View Article: PubMed Central - PubMed

Affiliation: Department of Anesthesiology and Pain Medicine, Center for Neuroscience, School of Medicine, University of California, Davis Davis, CA, USA.

ABSTRACT
As a result of cell-specific functions of voltage-activated K(+) channels, such as Kv7.1, mutations in this channel produce profound cardiac and auditory defects. At the same time, the massive diversity of K(+) channels allows for compensatory substitution of mutant channels by other functional channels of their type to minimize defective phenotypes. Kv7.1 represents a clear example of such functional dichotomy. While several point mutations in the channel result in a cardio-auditory syndrome called Jervell and Lange-Nielsen syndrome (JLNS), about 100-fold mutations result in long QT syndrome (LQTS) denoted as Romano-Ward syndrome (RWS), which has an intact auditory phenotype. To determine whether the cellular mechanisms for the diverse phenotypic outcome of Kv7.1 mutations, are dependent on the tissue-specific function of the channel and/or specialized functions of the channel, we made series of point mutations in hKv7.1 ascribed to JLNS and RWS. For JLNS mutations, all except W248F yielded non-functional channels when expressed alone. Although W248F at the end of the S4 domain yielded a functional current, it underwent marked inactivation at positive voltages, rendering the channel non-functional. We demonstrate that by definition, none of the JLNS mutants operated in a dominant negative (DN) fashion. Instead, the JLNS mutants have impaired membrane trafficking, trapped in the endoplasmic reticulum (ER) and Cis-Golgi. The RWS mutants exhibited varied functional phenotypes. However, they can be summed up as exhibiting DN effects. Phenotypic differences between JLNS and RWS may stem from tissue-specific functional requirements of cardiac vs. inner ear non-sensory cells.

No MeSH data available.


Related in: MedlinePlus