Large T-antigen up-regulates Kv4.3 K⁺ channels through Sp1, and Kv4.3 K⁺ channels contribute to cell apoptosis and necrosis through activation of calcium/calmodulin-dependent protein kinase II.
Down-regulation of Kv4.3 K⁺ channels commonly occurs in multiple diseases, but the understanding of the regulation of Kv4.3 K⁺ channels and the role of Kv4.3 K⁺ channels in pathological conditions are limited. HEK (human embryonic kidney)-293T cells are derived from HEK-293 cells which are transformed by expression of the large T-antigen. In the present study, by comparing HEK-293 and HEK-293T cells, we find that HEK-293T cells express more Kv4.3 K⁺ channels and more transcription factor Sp1 (specificity protein 1) than HEK-293 cells. Inhibition of Sp1 with Sp1 decoy oligonucleotide reduces Kv4.3 K⁺ channel expression in HEK-293T cells. Transfection of pN3-Sp1FL vector increases Sp1 protein expression and results in increased Kv4.3 K⁺ expression in HEK-293 cells. Since the ultimate determinant of the phenotype difference between HEK-293 and HEK-293T cells is the large T-antigen, we conclude that the large T-antigen up-regulates Kv4.3 K⁺ channel expression through an increase in Sp1. In both HEK-293 and HEK-293T cells, inhibition of Kv4.3 K⁺ channels with 4-AP (4-aminopyridine) or Kv4.3 small interfering RNA induces cell apoptosis and necrosis, which are completely rescued by the specific CaMKII (calcium/calmodulin-dependent protein kinase II) inhibitor KN-93, suggesting that Kv4.3 K⁺ channels contribute to cell apoptosis and necrosis through CaMKII activation. In summary, we establish: (i) the HEK-293 and HEK-293T cell model for Kv4.3 K⁺ channel study; (ii) that large T-antigen up-regulates Kv4.3 K⁺ channels through increasing Sp1 levels; and (iii) that Kv4.3 K⁺ channels contribute to cell apoptosis and necrosis through activating CaMKII. The present study provides deep insights into the mechanism of the regulation of Kv4.3 K⁺ channels and the role of Kv4.3 K⁺ channels in cell death.
Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), Harbin Medical University, Harbin 150086, People's Republic of China.
Qi
Q
Li
Ying
Y
Zhang
Yue
Y
Sheng
Rong
R
Huo
Bo
B
Sun
Xue
X
Teng
Na
N
Li
Jiu-Xin
JX
Zhu
Bao-Feng
BF
Yang
De-Li
DL
Dong
D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000378:metabolism* / Q000502:physiology; D049109:Cell Proliferation / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D005786:Gene Expression Regulation / Q000187:drug effects; D057809:HEK293 Cells; D006801:Humans; D008954:Models, Biological; D009336:Necrosis / Q000235:genetics* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D051672:Shal Potassium Channels / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D016329:Sp1 Transcription Factor / Q000502:physiology*; D015854:Up-Regulation / Q000235:genetics
D000957:Antigens, Viral, Tumor; D034741:RNA, Small Interfering; D051672:Shal Potassium Channels; D016329:Sp1 Transcription Factor; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2