Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na<sup>+</sup> Channel 1.2 Macromolecular Complex
Voltage-gated Na<sup>+</sup> (Na<sub>v</sub>) 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 Na<sub>v</sub...
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MDPI AG
2021-11-01
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| author | Nolan M. Dvorak Cynthia M. Tapia Timothy J. Baumgartner Jully Singh Fernanda Laezza Aditya K. Singh |
| author_facet | Nolan M. Dvorak Cynthia M. Tapia Timothy J. Baumgartner Jully Singh Fernanda Laezza Aditya K. Singh |
| author_sort | Nolan M. Dvorak |
| collection | DOAJ |
| description | Voltage-gated Na<sup>+</sup> (Na<sub>v</sub>) 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 Na<sub>v</sub> 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 Na<sub>v</sub> channel isoforms 1.2 (Na<sub>v</sub>1.2) and 1.6 (Na<sub>v</sub>1.6) are enriched, and their activities are differentially regulated by the Na<sub>v</sub> channel auxiliary protein fibroblast growth factor 14 (FGF14). Despite the known regulation of neuronal Na<sub>v</sub> 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 Na<sub>v</sub> 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 Na<sub>v</sub>1.2 and Na<sub>v</sub>1.6 channel macromolecular complexes. Our results show that the highly selective inhibitor of Wee1 kinase, called Wee1 inhibitor II, modulates FGF14:Na<sub>v</sub>1.2 complex assembly, but does not significantly affect FGF14:Na<sub>v</sub>1.6 complex assembly. These results are functionally recapitulated, as Wee1 inhibitor II entirely alters FGF14-mediated regulation of the Na<sub>v</sub>1.2 channel, but displays no effects on the Na<sub>v</sub>1.6 channel. At the molecular level, these effects of Wee1 inhibitor II on FGF14:Na<sub>v</sub>1.2 complex assembly and FGF14-mediated regulation of Na<sub>v</sub>1.2-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 Na<sub>v</sub>1.2 channel activity, which has important implications for unraveling cellular signaling pathways that fine-tune neuronal excitability. |
| first_indexed | 2024-03-10T05:36:53Z |
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| last_indexed | 2024-03-10T05:36:53Z |
| publishDate | 2021-11-01 |
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| spelling | doaj.art-f33551dbbfaa43bc9ba2d2846320a3bc2023-11-22T22:51:21ZengMDPI AGCells2073-44092021-11-011011310310.3390/cells10113103Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na<sup>+</sup> Channel 1.2 Macromolecular ComplexNolan M. Dvorak0Cynthia M. Tapia1Timothy J. Baumgartner2Jully Singh3Fernanda Laezza4Aditya K. Singh5Department of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, TX 75901, USADepartment of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, TX 75901, USADepartment of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, TX 75901, USADepartment of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, TX 75901, USADepartment of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, TX 75901, USADepartment of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, TX 75901, USAVoltage-gated Na<sup>+</sup> (Na<sub>v</sub>) 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 Na<sub>v</sub> 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 Na<sub>v</sub> channel isoforms 1.2 (Na<sub>v</sub>1.2) and 1.6 (Na<sub>v</sub>1.6) are enriched, and their activities are differentially regulated by the Na<sub>v</sub> channel auxiliary protein fibroblast growth factor 14 (FGF14). Despite the known regulation of neuronal Na<sub>v</sub> 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 Na<sub>v</sub> 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 Na<sub>v</sub>1.2 and Na<sub>v</sub>1.6 channel macromolecular complexes. Our results show that the highly selective inhibitor of Wee1 kinase, called Wee1 inhibitor II, modulates FGF14:Na<sub>v</sub>1.2 complex assembly, but does not significantly affect FGF14:Na<sub>v</sub>1.6 complex assembly. These results are functionally recapitulated, as Wee1 inhibitor II entirely alters FGF14-mediated regulation of the Na<sub>v</sub>1.2 channel, but displays no effects on the Na<sub>v</sub>1.6 channel. At the molecular level, these effects of Wee1 inhibitor II on FGF14:Na<sub>v</sub>1.2 complex assembly and FGF14-mediated regulation of Na<sub>v</sub>1.2-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 Na<sub>v</sub>1.2 channel activity, which has important implications for unraveling cellular signaling pathways that fine-tune neuronal excitability.https://www.mdpi.com/2073-4409/10/11/3103voltage-gated Na<sup>+</sup> (Na<sub>v</sub>) channelsfibroblast growth factor 14 (FGF14)Wee1 kinasepatch-clamp electrophysiology |
| spellingShingle | Nolan M. Dvorak Cynthia M. Tapia Timothy J. Baumgartner Jully Singh Fernanda Laezza Aditya K. Singh Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na<sup>+</sup> Channel 1.2 Macromolecular Complex Cells voltage-gated Na<sup>+</sup> (Na<sub>v</sub>) channels fibroblast growth factor 14 (FGF14) Wee1 kinase patch-clamp electrophysiology |
| title | Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na<sup>+</sup> Channel 1.2 Macromolecular Complex |
| title_full | Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na<sup>+</sup> Channel 1.2 Macromolecular Complex |
| title_fullStr | Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na<sup>+</sup> Channel 1.2 Macromolecular Complex |
| title_full_unstemmed | Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na<sup>+</sup> Channel 1.2 Macromolecular Complex |
| title_short | Pharmacological Inhibition of Wee1 Kinase Selectively Modulates the Voltage-Gated Na<sup>+</sup> Channel 1.2 Macromolecular Complex |
| title_sort | pharmacological inhibition of wee1 kinase selectively modulates the voltage gated na sup sup channel 1 2 macromolecular complex |
| topic | voltage-gated Na<sup>+</sup> (Na<sub>v</sub>) channels fibroblast growth factor 14 (FGF14) Wee1 kinase patch-clamp electrophysiology |
| url | https://www.mdpi.com/2073-4409/10/11/3103 |
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