The TMEM16A Channel Mediates the fast polyspermy block in Xenopus Laevis
| Autor: | Anne E. Carlson, Wesley A. Phelps, Miler T. Lee, Katherine L. Wozniak, Maiwase Tembo |
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| Rok vydání: | 2018 |
| Předmět: |
Male
0301 basic medicine African clawed frog Physiology Xenopus Xenopus laevis 03 medical and health sciences 0302 clinical medicine Human fertilization medicine Animals Research Articles 030304 developmental biology 0303 health sciences biology Chemistry Embryo Depolarization biology.organism_classification Polyspermy Spermatozoa Sperm Cell biology 030104 developmental biology Bestrophin 1 medicine.anatomical_structure biology.protein Gamete Calcium Developmental biology 030217 neurology & neurosurgery Research Article |
| Zdroj: | The Journal of General Physiology |
| Popis: | In their preceding paper, Wozniak et al. show that fertilization increases intracellular Ca2+ in Xenopus laevis eggs by activating an IP3 signaling cascade. Here, they reveal that Ca2+ subsequently opens the Cl− channel TMEM16A to allow Cl− efflux, cell depolarization, and fast block to polyspermy. In externally fertilizing animals, such as sea urchins and frogs, prolonged depolarization of the egg immediately after fertilization inhibits the entry of additional sperm—a phenomenon known as the fast block to polyspermy. In the African clawed frog Xenopus laevis, this depolarization is driven by Ca2+-activated Cl− efflux. Although the prominent Ca2+-activated Cl− currents generated in immature X. laevis oocytes are mediated by X. laevis transmembrane protein 16a (xTMEM16A) channels, little is known about the channels that contribute to the fast block in mature eggs. Moreover, the gamete undergoes a gross transformation as it develops from an immature oocyte into a fertilization-competent egg. Here, we report the results of our approach to identify the Ca2+-activated Cl− channel that triggers the fast block. By querying published proteomic and RNA-sequencing data, we identify two Ca2+-activated Cl− channels expressed in fertilization-competent X. laevis eggs: xTMEM16A and X. laevis bestrophin 2A (xBEST2A). By exogenously expressing xTMEM16A and xBEST2A in axolotl cells lacking endogenous Ca2+-activated currents, we characterize the effect of inhibitors on currents mediated by these channels. None of the inhibitors tested block xBEST2A currents specifically. However, 2-(4-chloro-2-methylphenoxy)-N-[(2-methoxyphenyl)methylideneamino]-acetamide (Ani9) and N-((4-methoxy)-2-naphthyl)-5-nitroanthranilic acid (MONNA) each reduce xTMEM16A currents by more than 70% while only nominally inhibiting those generated by xBEST2A. Using whole-cell recordings during fertilization, we find that Ani9 and MONNA effectively diminish fertilization-evoked depolarizations. Additionally, these inhibitors lead to increased polyspermy in X. laevis embryos. These results indicate that fertilization activates TMEM16A channels in X. laevis eggs and induces the earliest known event triggered by fertilization: the fast block to polyspermy. |
| Databáze: | OpenAIRE |
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