scholarly journals Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na+ Channel 1.2 Macromolecular Complex

Cells ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 3103
Author(s):  
Nolan M. Dvorak ◽  
Cynthia M. Tapia ◽  
Timothy J. Baumgartner ◽  
Jully Singh ◽  
Fernanda Laezza ◽  
...  

Voltage-gated Na+ (Nav) channels are a primary molecular determinant of the action potential (AP). Despite the canonical role of the pore-forming α subunit in conferring this function, protein–protein interactions (PPI) between the Nav channel α subunit and its auxiliary proteins are necessary to reconstitute the full physiological activity of the channel and to fine-tune neuronal excitability. In the brain, the Nav channel isoforms 1.2 (Nav1.2) and 1.6 (Nav1.6) are enriched, and their activities are differentially regulated by the Nav channel auxiliary protein fibroblast growth factor 14 (FGF14). Despite the known regulation of neuronal Nav channel activity by FGF14, less is known about cellular signaling molecules that might modulate these regulatory effects of FGF14. To that end, and building upon our previous investigations suggesting that neuronal Nav channel activity is regulated by a kinase network involving GSK3, AKT, and Wee1, we interrogate in our current investigation how pharmacological inhibition of Wee1 kinase, a serine/threonine and tyrosine kinase that is a crucial component of the G2-M cell cycle checkpoint, affects the Nav1.2 and Nav1.6 channel macromolecular complexes. Our results show that the highly selective inhibitor of Wee1 kinase, called Wee1 inhibitor II, modulates FGF14:Nav1.2 complex assembly, but does not significantly affect FGF14:Nav1.6 complex assembly. These results are functionally recapitulated, as Wee1 inhibitor II entirely alters FGF14-mediated regulation of the Nav1.2 channel, but displays no effects on the Nav1.6 channel. At the molecular level, these effects of Wee1 inhibitor II on FGF14:Nav1.2 complex assembly and FGF14-mediated regulation of Nav1.2-mediated Na+ currents are shown to be dependent upon the presence of Y158 of FGF14, a residue known to be a prominent site for phosphorylation-mediated regulation of the protein. Overall, our data suggest that pharmacological inhibition of Wee1 confers selective modulatory effects on Nav1.2 channel activity, which has important implications for unraveling cellular signaling pathways that fine-tune neuronal excitability.

2013 ◽  
Vol 305 (4) ◽  
pp. H431-H445 ◽  
Author(s):  
Anthony W. Herren ◽  
Donald M. Bers ◽  
Eleonora Grandi

The voltage-gated Na channel isoform 1.5 (NaV1.5) is the pore forming α-subunit of the voltage-gated cardiac Na channel, which is responsible for the initiation and propagation of cardiac action potentials. Mutations in the SCN5A gene encoding NaV1.5 have been linked to changes in the Na current leading to a variety of arrhythmogenic phenotypes, and alterations in the NaV1.5 expression level, Na current density, and/or gating have been observed in acquired cardiac disorders, including heart failure. The precise mechanisms underlying these abnormalities have not been fully elucidated. However, several recent studies have made it clear that NaV1.5 forms a macromolecular complex with a number of proteins that modulate its expression levels, localization, and gating and is the target of extensive post-translational modifications, which may also influence all these properties. We review here the molecular aspects of cardiac Na channel regulation and their functional consequences. In particular, we focus on the molecular and functional aspects of Na channel phosphorylation by the Ca/calmodulin-dependent protein kinase II, which is hyperactive in heart failure and has been causally linked to cardiac arrhythmia. Understanding the mechanisms of altered NaV1.5 expression and function is crucial for gaining insight into arrhythmogenesis and developing novel therapeutic strategies.


2021 ◽  
Vol 8 ◽  
Author(s):  
Aditya K. Singh ◽  
Nolan M. Dvorak ◽  
Cynthia M. Tapia ◽  
Angela Mosebarger ◽  
Syed R. Ali ◽  
...  

The voltage-gated Na+ (Nav) channel is a primary molecular determinant of the initiation and propagation of the action potential. Despite the central role of the pore-forming α subunit in conferring this functionality, protein:protein interactions (PPI) between the α subunit and auxiliary proteins are necessary for the full physiological activity of Nav channels. In the central nervous system (CNS), one such PPI occurs between the C-terminal domain of the Nav1.6 channel and fibroblast growth factor 14 (FGF14). Given the primacy of this PPI in regulating the excitability of neurons in clinically relevant brain regions, peptides targeting the FGF14:Nav1.6 PPI interface could be of pre-clinical value. In this work, we pharmacologically evaluated peptides derived from FGF14 that correspond to residues that are at FGF14’s PPI interface with the CTD of Nav1.6. These peptides, Pro-Leu-Glu-Val (PLEV) and Glu-Tyr-Tyr-Val (EYYV), which correspond to residues of the β12 sheet and β8-β9 loop of FGF14, respectively, were shown to inhibit FGF14:Nav1.6 complex assembly. In functional studies using whole-cell patch-clamp electrophysiology, PLEV and EYYV were shown to confer differential modulation of Nav1.6-mediated currents through mechanisms dependent upon the presence of FGF14. Crucially, these FGF14-dependent effects of PLEV and EYYV on Nav1.6-mediated currents were further shown to be dependent on the N-terminal domain of FGF14. Overall, these data suggest that the PLEV and EYYV peptides represent scaffolds to interrogate the Nav1.6 channel macromolecular complex in an effort to develop targeted pharmacological modulators.


2005 ◽  
Vol 26 (5) ◽  
pp. 743-752 ◽  
Author(s):  
Hiroaki Misonou ◽  
Durga P. Mohapatra ◽  
James S. Trimmer

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Nina E Ottosson ◽  
Xiongyu Wu ◽  
Andreas Nolting ◽  
Urban Karlsson ◽  
Per-Eric Lund ◽  
...  

2007 ◽  
Vol 35 (5) ◽  
pp. 1064-1068 ◽  
Author(s):  
D.P. Mohapatra ◽  
K.-S. Park ◽  
J.S. Trimmer

Voltage-gated K+ channels are key regulators of neuronal excitability. The Kv2.1 voltage-gated K+ channel is the major delayed rectifier K+ channel expressed in most central neurons, where it exists as a highly phosphorylated protein. Kv2.1 plays a critical role in homoeostatic regulation of intrinsic neuronal excitability through its activity- and calcineurin-dependent dephosphorylation. Here, we review studies leading to the identification and functional characterization of in vivo Kv2.1 phosphorylation sites, a subset of which contribute to graded modulation of voltage-dependent gating. These findings show that distinct developmental-, cell- and state-specific regulation of phosphorylation at specific sites confers a diversity of functions on Kv2.1 that is critical to its role as a regulator of intrinsic neuronal excitability.


2018 ◽  
Vol 137 (1) ◽  
pp. 93-97 ◽  
Author(s):  
Takafumi Horishita ◽  
Nobuyuki Yanagihara ◽  
Susumu Ueno ◽  
Dan Okura ◽  
Reiko Horishita ◽  
...  

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