Phosphatidylinositol 4,5-bisphosphate (PIP(2)) and Ca(2+) are both required to open the Cl(−) channel TMEM16A
| Autor: | Rachel E. Bainbridge, Maiwase Tembo, Anne E. Carlson, Katherine L. Wozniak |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
Phosphatidylinositol 4 5-Diphosphate Xenopus Gating Biochemistry Membrane Potentials 03 medical and health sciences chemistry.chemical_compound Xenopus laevis Chloride Channels Membrane Biology Animals Phosphatidylinositol Patch clamp Calcium Signaling Molecular Biology 030102 biochemistry & molecular biology biology Cell Biology Smooth muscle contraction biology.organism_classification Cell biology 030104 developmental biology Phosphatidylinositol 4 5-bisphosphate chemistry Second messenger system Chloride channel lipids (amino acids peptides and proteins) Calcium Ion Channel Gating |
| Zdroj: | J Biol Chem |
| Popis: | Transmembrane member 16A (TMEM16A) is a widely expressed Ca(2+)-activated Cl(−) channel with various physiological functions ranging from mucosal secretion to regulating smooth muscle contraction. Understanding how TMEM16A controls these physiological processes and how its dysregulation may cause disease requires a detailed understanding of how cellular processes and second messengers alter TMEM16A channel gating. Here we assessed the regulation of TMEM16A gating by recording Ca(2+)-evoked Cl(−) currents conducted by endogenous TMEM16A channels expressed in Xenopus laevis oocytes, using the inside-out configuration of the patch clamp technique. During continuous application of Ca(2+), we found that TMEM16A-conducted currents decay shortly after patch excision. Such current rundown is common among channels regulated by phosphatidylinositol 4,5-bisphosphate (PIP(2)). Thus, we sought to investigate a possible role of PIP(2) in TMEM16A gating. Consistently, synthetic PIP(2) rescued the current after rundown, and the application of PIP(2) modulating agents altered the speed kinetics of TMEM16A current rundown. First, two PIP(2) sequestering agents, neomycin and anti-PIP(2), applied to the intracellular surface of excised patches sped up TMEM16A current rundown to nearly twice as fast. Conversely, rephosphorylation of phosphatidylinositol (PI) derivatives into PIP(2) using Mg-ATP or inhibiting dephosphorylation of PIP(2) using β-glycerophosphate slowed rundown by nearly 3-fold. Our results reveal that TMEM16A regulation is more complicated than it initially appeared; not only is Ca(2+) necessary to signal TMEM16a opening, but PIP(2) is also required. These findings improve our understanding of how the dysregulation of these pathways may lead to disease and suggest that targeting these pathways could have utility for potential therapies. |
| Databáze: | OpenAIRE |
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