14-3-3 promotes sarcolemmal expression of cardiac CaV1.2 and nucleates isoproterenol-triggered channel superclustering

• 成果类型: Article
• 署名作者: Spooner, Heather C.; Costa, Alexandre D.; Westhoff, Maartje; Hernandez-Gonzalez, Adriana; Ibrahimkhail, Husna; Yarov-Yarovoy, Vladimir; Horne, Mary C.; Dickson, Eamonn J.; Dixon, Rose E.
• 署名单位: University of California System; University of California Davis; University of California System; University of California Davis; University of California System; University of California Davis
• 刊物名称: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• ISSN/ISSBN: 0027-12484
• DOI: 10.1073/pnas.241330812
• 发表日期: 2025-02-04
• 关键词: inhibitory interaction; surface expression; calcium-channels; beta-subunits; endoplasmic-reticulum; protein-kinase; binding; phosphorylation; heart; rad

摘要

The L- type Ca2+ channel (Ca(V)1.2) is essential for cardiac excitation-contraction coupling. To contribute to the inward Ca2+ flux that drives Ca2+- induced- Ca2+- release, Ca(V)1.2 channels must be expressed on the sarcolemma; thus the regulatory mechanisms that tune Ca(V)1.2 expression to meet contractile demand are an emerging area of research. A ubiquitously expressed protein called 14- 3- 3 has been proposed to affect Ca2+ channel trafficking in nonmyocytes; however, whether 14- 3- 3 has similar effects on Ca(V)1.2 in car-diomyocytes is unknown. 14- 3- 3 preferentially binds phospho- serine/threonine residues to affect many cellular processes and is known to regulate cardiac ion channels including NaV1.5 and the human ether- & agrave;- go- go-related gene (hERG) potassium channel. Altered 14- 3- 3 expression and function have been implicated in cardiac pathologies including hypertrophy. Accordingly, we tested the hypothesis that 14- 3- 3 interacts with Ca(V)1.2 in a phosphorylation- dependent manner and regulates cardiac Ca(V)1.2 trafficking and recy-cling. Confocal imaging, proximity ligation assays, superresolution imaging, and coim-munoprecipitation revealed a population of 14- 3- 3 colocalized and closely associated with Ca(V)1.2. The degree of 14- 3- 3/Ca(V)1.2 colocalization increased upon stimulation of beta- adrenergic receptors with isoproterenol. Notably, only the 14- 3- 3- associated Ca(V)1.2 population displayed increased cluster size with isoproterenol, revealing a role for 14- 3- 3 as a nucleation factor that directs Ca(V)1.2 superclustering. Isoproterenol- stimulated augmentation of sarcolemmal Ca(V)1.2 expression, Ca2+ currents, and Ca2+ transients in ventricular myocytes were strengthened by 14- 3- 3 overexpression and attenuated by 14- 3- 3 inhibition. These data support a model where 14- 3- 3 interacts with Ca(V)1.2 in a phosphorylation- dependent manner to promote enhanced trafficking/recycling, clustering, and activity during beta- adrenergic stimulation.