Two arginines in the cytoplasmic C-terminal domain are essential for voltage-dependent regulation of A-type K+ current in the Kv4 channel subfamily.
Contributions of the C-terminal domain of Kv4.3 to the voltage-dependent gating of A-type K+ current (IA) were examined by (i) making mutations in this region, (ii) heterologous expression in HEK293 cells, and (iii) detailed voltage clamp analyses. Progressive deletions of the C terminus of rat Kv4.3M (to amino acid 429 from the N terminus) did not markedly change the inactivation time course of IA but shifted the voltage dependence of steady state inactivation in the negative direction to a maximum of -17 mV. Further deletions (to amino acid 420) shifted this parameter in the positive direction, suggesting a critical role for the domain 429-420 in the voltage-dependent regulation of IA. There are four positively charged amino acids in this domain: Lys423, Lys424, Arg426, and Arg429. The replacement of the two arginines with alanines (R2A) resulted in -23 and -13 mV shifts of inactivation and activation, respectively. Additional replacement of the two lysines with alanines did not result in further shifts. Single replacements of R426A or R429A induced -15 and -10 mV shifts of inactivation, respectively. R2A did not significantly change the inactivation rate but did markedly change the voltage dependence of recovery from inactivation. These two arginines are conserved in Kv4 subfamily, and alanine replacement of Arg429 and Arg432 in Kv4.2 gave essentially the same results. These effects of R2A were not modulated by co-expression of the K+ channel beta subunit, KChIPs. In conclusion, the two arginines in the cytosolic C-terminal domain of alpha-subunits of Kv4 subfamily strongly regulate the voltage dependence of channel activation, inactivation, and recovery.
Hatano
Noriyuki
N
Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan.
Ohya
Susumu
S
Muraki
Katsuhiko
K
Clark
Robert B
RB
Giles
Wayne R
WR
Imaizumi
Yuji
Y
D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D001120:Arginine / Q000737:chemistry*; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D017353:Gene Deletion; D006801:Humans; D007700:Kinetics; D008239:Lysine / Q000737:chemistry; D008969:Molecular Sequence Data; D009154:Mutation; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000737:chemistry* / Q000235:genetics; D024642:Potassium Channels, Voltage-Gated; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D051672:Shal Potassium Channels; D013997:Time Factors; D014162:Transfection
D018076:DNA, Complementary; C497018:KCND2 protein, human; C497021:KCND3 protein, human; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D051672:Shal Potassium Channels; D001120:Arginine; D008239:Lysine; D000409:Alanine