Emerging Concepts in Ion Channel Biophysics

Emerging Concepts in Ion Channel Biophysics

Poster Abstracts

4-POS Board 4 Functional Analysis of the Voltage Sensor Domain Present in the Mammalian Sperm- Specific Na + /H + Exchanger by Patch-Clamp Fluorometry of Chimeric Fluorescent Voltage Sensor César Arcos Hernández , Takuya Nishigaki. Instituto de Biotecnología, Cuernavaca, Morelos, Mexico. Sperm motility is accurately regulated by several enzymes, transporters and ion channels. Some of these proteins are expressed only in spermatozoa, but the functional relationship among them is not fully understood. Sperm-specific Na + /H + exchanger (sNHE) is an essential protein for mouse sperm motility regulation and is localized in the principal piece of mammalian spermatozoa. Differently from somatic NHEs, sNHE has two predicted regulatory domains, a voltage sensor domain (VSD) and a cyclic nucleotide binding domain (CNBD), although their functionality remains to be confirmed. In the case of the VSD, it has been proposed that hyperpolarization caused by Slo3 (a sperm specific K + channel) promotes an intracellular alkalization by the sNHE through this VSD. This signal cascade is thought to be involved in sperm hyperactivation. Interestingly, there are experiments that suggest the VSD of sNHE is functional in mice but not in humans. Correlating this, the fourth transmembrane segment of the VSD of the human sNHE, but not the mouse sNHE, lacks an arginine in a critical position, which could cause the domain not to be functional. To demonstrate functional differences in the VSD of the sNHE between human and mouse, we will produce fluorescent voltage sensors (VSFPs) using Arclight, a popular VSFP, as a template and replace its VSD by that of sNHE. We will analyze the biophysical properties of the VSFPs expressed in HEK293 cells by using patch clamp fluorometry and extrapolate the function of the VSDs of human and mouse sNHE. This project is supported by PAPIIT (DGAPA IN206116).

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