Limits...
Regulated RNA editing and functional epistasis in Shaker potassium channels.

Ingleby L, Maloney R, Jepson J, Horn R, Reenan R - J. Gen. Physiol. (2009)

Bottom Line: Genetic manipulations of editing enzyme activity demonstrated that a chief determinant of Shaker editing site choice resides not in the editing enzyme, but rather, in unknown factors intrinsic to cells.Characterizing the biophysical properties of currents in nine isoforms revealed an unprecedented feature, functional epistasis; biophysical phenotypes of isoforms cannot be explained simply by the consequences of individual editing effects at the four sites.Our results unmask allosteric communication across disparate regions of the channel protein and between evolved and regulated amino acid changes introduced by RNA editing.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Physiology and Biophysics, Institute of Hyperexcitability, Jefferson Medical College, Philadelphia, PA 19107, USA.

ABSTRACT
Regulated point modification by an RNA editing enzyme occurs at four conserved sites in the Drosophila Shaker potassium channel. Single mRNA molecules can potentially represent any of 2(4) = 16 permutations (isoforms) of these natural variants. We generated isoform expression profiles to assess sexually dimorphic, spatial, and temporal differences. Striking tissue-specific expression was seen for particular isoforms. Moreover, isoform distributions showed evidence for coupling (linkage) of editing sites. Genetic manipulations of editing enzyme activity demonstrated that a chief determinant of Shaker editing site choice resides not in the editing enzyme, but rather, in unknown factors intrinsic to cells. Characterizing the biophysical properties of currents in nine isoforms revealed an unprecedented feature, functional epistasis; biophysical phenotypes of isoforms cannot be explained simply by the consequences of individual editing effects at the four sites. Our results unmask allosteric communication across disparate regions of the channel protein and between evolved and regulated amino acid changes introduced by RNA editing.

Show MeSH
Kinetics of deactivation and inactivation. (A) Deactivation kinetics in the two most extreme isoforms of inactivation-removed mutants. Tail current kinetics measured over a range from −60 to −140 mV (Table S4). (B) Poor correlation between inactivation and deactivation time constants among the isoforms.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC2606942&req=5

fig8: Kinetics of deactivation and inactivation. (A) Deactivation kinetics in the two most extreme isoforms of inactivation-removed mutants. Tail current kinetics measured over a range from −60 to −140 mV (Table S4). (B) Poor correlation between inactivation and deactivation time constants among the isoforms.

Mentions: A noticeable difference among the isoforms shown in Fig. 6 is that AAGA has faster kinetics of deactivation at −120 mV than those of the other three isoforms. The rate of deactivation varied widely among the isoforms (Table S4). Fig. 8 (A and B) demonstrates this point with tail currents from the fastest (AAGG) and slowest (AGAA) deactivating isoforms over a voltage range of −60 to −140 mV. Fig. 8 C shows a complete lack of correlation between the kinetics of deactivation and those of inactivation. This suggests that the rate of ball-and-chain inactivation in these isoforms is controlled by factors quite separate from those controlling movement of the activation gate. The kinetics of gate opening were not examined in detail because of the kinetic overlap of gating and ionic current under the conditions of these experiments.


Regulated RNA editing and functional epistasis in Shaker potassium channels.

Ingleby L, Maloney R, Jepson J, Horn R, Reenan R - J. Gen. Physiol. (2009)

Kinetics of deactivation and inactivation. (A) Deactivation kinetics in the two most extreme isoforms of inactivation-removed mutants. Tail current kinetics measured over a range from −60 to −140 mV (Table S4). (B) Poor correlation between inactivation and deactivation time constants among the isoforms.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2606942&req=5

fig8: Kinetics of deactivation and inactivation. (A) Deactivation kinetics in the two most extreme isoforms of inactivation-removed mutants. Tail current kinetics measured over a range from −60 to −140 mV (Table S4). (B) Poor correlation between inactivation and deactivation time constants among the isoforms.
Mentions: A noticeable difference among the isoforms shown in Fig. 6 is that AAGA has faster kinetics of deactivation at −120 mV than those of the other three isoforms. The rate of deactivation varied widely among the isoforms (Table S4). Fig. 8 (A and B) demonstrates this point with tail currents from the fastest (AAGG) and slowest (AGAA) deactivating isoforms over a voltage range of −60 to −140 mV. Fig. 8 C shows a complete lack of correlation between the kinetics of deactivation and those of inactivation. This suggests that the rate of ball-and-chain inactivation in these isoforms is controlled by factors quite separate from those controlling movement of the activation gate. The kinetics of gate opening were not examined in detail because of the kinetic overlap of gating and ionic current under the conditions of these experiments.

Bottom Line: Genetic manipulations of editing enzyme activity demonstrated that a chief determinant of Shaker editing site choice resides not in the editing enzyme, but rather, in unknown factors intrinsic to cells.Characterizing the biophysical properties of currents in nine isoforms revealed an unprecedented feature, functional epistasis; biophysical phenotypes of isoforms cannot be explained simply by the consequences of individual editing effects at the four sites.Our results unmask allosteric communication across disparate regions of the channel protein and between evolved and regulated amino acid changes introduced by RNA editing.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Physiology and Biophysics, Institute of Hyperexcitability, Jefferson Medical College, Philadelphia, PA 19107, USA.

ABSTRACT
Regulated point modification by an RNA editing enzyme occurs at four conserved sites in the Drosophila Shaker potassium channel. Single mRNA molecules can potentially represent any of 2(4) = 16 permutations (isoforms) of these natural variants. We generated isoform expression profiles to assess sexually dimorphic, spatial, and temporal differences. Striking tissue-specific expression was seen for particular isoforms. Moreover, isoform distributions showed evidence for coupling (linkage) of editing sites. Genetic manipulations of editing enzyme activity demonstrated that a chief determinant of Shaker editing site choice resides not in the editing enzyme, but rather, in unknown factors intrinsic to cells. Characterizing the biophysical properties of currents in nine isoforms revealed an unprecedented feature, functional epistasis; biophysical phenotypes of isoforms cannot be explained simply by the consequences of individual editing effects at the four sites. Our results unmask allosteric communication across disparate regions of the channel protein and between evolved and regulated amino acid changes introduced by RNA editing.

Show MeSH