Formation of Heterotetrameric Potassium Channels Kv1.1–Kv1.2 in Neuro-2a Cells: Analysis by the Förster Resonance Energy Transfer Technique

Cover Page

Cite item

Full Text

Open Access Open Access
Restricted Access Access granted
Restricted Access Subscription Access

Abstract

The subunits of the voltage-gated potassium channels Kv1.1 and Kv1.2 can form both homo- and heterotetrameric channels in cells. This significantly affects the functional properties and localization of the formed Kv1 channels. Confocal microscopy based on Förster resonance energy transfer was used to study the formation of Kv1 channels during co-expression of subunits Kv1.1(S369T) and Kv1.2(S371T), fused with the fluorescent protein mKate2 and TagCFP, respectively, in murine neuroblastoma Neuro-2a cells. Due to mutation, these subunits provide enhanced transfer of Kv1 channels in plasma membrane. It was found that TagCFP-Kv1.1(S369T) and mKate2-Kv1.2(S371T) effectively form heterochannels that are localized both on the membrane and in the cytoplasm of cells. In the absence of the S369T mutation, heterochannels are not embedded in the membrane, which indicates the need for auxiliary factors for the transfer of native heterochannels into the cell membrane. In addition to heterochannels, homotetrameric channels are also formed in cells, but the effectiveness of the formation of heterochannels is much higher.

About the authors

A. V Efremenko

Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences

Moscow, Russia

O. V Nekrasova

Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences

Moscow, Russia

A. V Feofanov

Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences; Lomonosov Moscow State University

Email: avfeofanov@yandex.ru
Moscow, Russia; Moscow, Russia

References

  1. Ovsepian S. V., Leberre M., Steuber V., O'Leary V. B., Leibold Ch. J., and Dolly O. Distinctive role of Kv1.1 subunit in the biology and functions of low threshold K+ channels with implications for neurological disease. Pharmacol. Ther., 159, 93–101 (2016). doi: 10.1016/j.pharmthera.2016.01.005
  2. Al-Sabi A., Kaza S. K., Dolly O. J., and Wang J. Pharmacological characteristics of Kv1.1- and Kv1.2-containing channels are influenced by the stoichiometry and positioning of their α subunits. Biochem. J., 454, 101–108 (2013). doi: 10.1042/BJ20130297
  3. Robbins C. A. and Tempel B. L. Kv1.1 and Kv1.2: similar channels, different seizure models. Epilepsia, 53 (Suppl 1), 134–141 (2012). doi: 10.1111/J.1528-1167.2012.03484.X
  4. Ma Z., Lavebratt C., Almgren M., Portwood N., Forsberg L. E., Bränström R., Berglund E., Falkmer S., Sundler F., Wierup N., and Björklund A. Evidence for presence and functional effects of Kv1.1 channels in β-cells: general survey and results from mceph/mceph mice. PLoS One, 6 (4), e18213 (2011). doi: 10.1371/journal.pone.0018213
  5. Fellerhoff-Losch B., Korol S. V., Ganor Y., Gu S., Cooper I., Eilam R., Besser M., Goldfinger M., Chowers Y., Wank R., Birnir B., and Levite M. Normal human CD4(+) helper T cells express Kv1.1 voltage-gated K+ channels, and selective Kv1.1 block in T cells induces by it-self robust TNFα production and secretion and activation of the NFκB non-canonical pathway. J. Neural Transm.(Vienna), 123, 137–157 (2016). doi: 10.1007/s00702-015-1446-9
  6. Park W. S., Firth A. L., Han J., and Ko E. A. Patho-, physiological roles of voltage-dependent K+ channels in pulmonary arterial smooth muscle cells. J. Smooth Muscle Res., 46, 89–105 (2010). doi: 10.1540/jsmr.46.89
  7. Pinatel D. and Faivre-Sarrailh C. Assembly and Function of the Juxtaparanodal Kv1 Complex in Health and Disease. Life (Basel), 11, 1–22 (2020). doi: 10.3390/LIFE11010008
  8. Manganas L. N. and Trimmer J. S. Subunit composition determines Kv1 potassium channel surface expression. J. Biol. Chem., 275, 29685–29693 (2000). doi: 10.1074/JBC.M005010200
  9. Duménieu M., Oulé M., Kreutz M. R., and LopezRojas J. The segregated expression of voltage-gated potassium and sodium channels in neuronal membranes: functional implications and regulatory mechanisms. Front. Cell. Neurosci., 11, 115 (2017). doi: 10.3389/fncel.2017.00115
  10. Capera J., Serrano-Novillo C., Navarro-Pérez M., Cassinelli S., and Felipe A. The potassium channel odyssey: Mechanisms of traffic and membrane arrangement. Int. J. Mol. Sci., 20, 734 (2019). doi: 10.3390/ijms20030734
  11. Manganas L. N., Wang Q., Scannevin R. H., and Trimmer J. S. Identification of a trafficking determinant localized to the Kv1 potassium channel pore. Proc. Natl. Acad. Sci. USA, 98, 14055–14059 (2001). doi: 10.1073/PNAS.241403898
  12. Zhu J., Watanabe I., Gomez B., and Thornhill W. B. Determinants involved in Kv1 potassium channel folding in the endoplasmic reticulum, glycosylation in the Golgi, and cell surface expression. J. Biol. Chem., 276, 39419–39427 (2001). doi: 10.1074/JBC.M107399200
  13. Zhu J., Gomez B., Watanabe I., and Thornhill W. B. Amino acids in the pore region of Kv1 potassium channels dictate cell-surface protein levels: a possible trafficking code in the Kv1 subfamily. Biochem. J., 388, 355–362 (2005). doi: 10.1042/BJ20041447
  14. Orlov N. A., Kryukova E. V., Efremenko A. V., Yakimov S. A., Toporova V. A., Kirpichnikov M. P., Nekrasova O. V., and Feofanov A. V. Interactions of the Kv1.1 channel with peptide pore blockers: A fluorescent analysis on mammalian cells. Membranes (Basel), 13 (7), 645 (2023). doi: 10.3390/membranes13070645
  15. Ignatova A. A., Kryukova E. V., Novoseletsky V. N., Kazakov O. V., Orlov N. A., Korabeynikova V. N., Larina M. V., Fradkov A. F., Yakimov S. A., Kirpichnikov M. P., Feofanov A. V., and Nekrasova O. V. New high-affinity peptide ligands for Kv1.2 channel: selective blockers and fluorescent probes. Cells, 13 (24), 2096 (2024). doi: 10.3390/cells13242096
  16. Orlov N. A., Ignatova A. A., Kryukova E. V., Yakimov S. A., Kirpichnikov M. P., Nekrasova O. V., and Feofanov A. V. Combining mKate2-Kv1.3 channel and Atto488-hongotoxin for the studies of peptide pore blockers on living eukaryotic cells. Toxins (Basel), 14 (12), 858 (2022). doi: 10.3390/toxins14120858

Supplementary files

Supplementary Files
Action
1. JATS XML
Download

Copyright (c) 2025 Russian Academy of Sciences

Согласие на обработку персональных данных

 

Используя сайт https://journals.rcsi.science, я (далее – «Пользователь» или «Субъект персональных данных») даю согласие на обработку персональных данных на этом сайте (текст Согласия) и на обработку персональных данных с помощью сервиса «Яндекс.Метрика» (текст Согласия).