Functional expression of Kir2.x in human aortic endothelial cells: the dominant role of Kir2.2.
Inward rectifier K(+) channels (Kir) are a significant determinant of endothelial cell (EC) membrane potential, which plays an important role in endothelium-dependent vasodilatation. In the present study, several complementary strategies were applied to determine the Kir2 subunit composition of human aortic endothelial cells (HAECs). Expression levels of Kir2.1, Kir2.2, and Kir2.4 mRNA were similar, whereas Kir2.3 mRNA expression was significantly weaker. Western blot analysis showed clear Kir2.1 and Kir2.2 protein expression, but Kir2.3 protein was undetectable. Functional analysis of endothelial inward rectifier K(+) current (I(K)) demonstrated that 1) I(K) current sensitivity to Ba(2+) and pH were consistent with currents determined using Kir2.1 and Kir2.2 but not Kir2.3 and Kir2.4, and 2) unitary conductance distributions showed two prominent peaks corresponding to known unitary conductances of Kir2.1 and Kir2.2 channels with a ratio of approximately 4:6. When HAECs were transfected with dominant-negative (dn)Kir2.x mutants, endogenous current was reduced approximately 50% by dnKir2.1 and approximately 85% by dnKir2.2, whereas no significant effect was observed with dnKir2.3 or dnKir2.4. These studies suggest that Kir2.2 and Kir2.1 are primary determinants of endogenous K(+) conductance in HAECs under resting conditions and that Kir2.2 provides the dominant conductance in these cells.
Fang
Yun
Y
Institute for Medicine and Engineering, University of Pennsylvania, 1160 Vagelos Research Labs, 3340 Smith Walk, Philadelphia, PA 19104, USA.
Schram
Gernot
G
Romanenko
Victor G
VG
Shi
Congzhu
C
Conti
Lisa
L
Vandenberg
Carol A
CA
Davies
Peter F
PF
Nattel
Stanley
S
Levitan
Irena
I
D001011:Aorta / Q000166:cytology; D001464:Barium / Q000494:pharmacology; D002462:Cell Membrane / Q000502:physiology; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008564:Membrane Potentials / Q000502:physiology; D011188:Potassium / Q000378:metabolism; D024661:Potassium Channels, Inwardly Rectifying / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000096:biosynthesis
C099489:KCNJ4 protein, human; C489461:Kir2.1 channel; C489463:Kir2.2 channel; C489465:Kir2.4 channel; D024661:Potassium Channels, Inwardly Rectifying; D012333:RNA, Messenger; D001464:Barium; D011188:Potassium