FUNCTIONAL CHARACTERIZATION OF THE SLC26A9 ION CHANNEL PERMEATION PATHWAY

The long-term goal of this project is to develop a detailed molecular understanding of the SLC26A9 anion channel. SLC26A9 belongs to the SLC26 family of membrane transporters which are found in various organisms, from prokaryotes to mammals. The members of the SLC26 family play central roles in the secretion and absorption of nutrients and many anionic substrates such as SO42−, Cl−, HCO3− or I− which are essential for the cell's physiological processes (e.g. swelling and salt homeostasis). As with other members of its family, SLC26A9 has a modular architecture consisting of two halves, each possessing seven transmembrane segments followed by a cytoplasmic sulfate transporter and an antisigma factor antagonist (STAT) domain. At the functional level, studies have proposed that SLC26A9 functions as a Cl− /HCO3− transporter as well as a Cl− channel; therefore, SLC26A9 is thought to have opposite functional properties, mediating active and passive transport, respectively. These discrepancies in the function of SLC26A9 raise several significant questions about its actual functional mode. Our preliminary data demonstrates that SLC26A9 is a Cl− channel and our main goal is to characterise its function through the investigation of its transport mechanism. In this project, we also aim to identify the parts of the protein that form the Cl− transport pathway and the key structural features essential to support its function as a channel rather than a transporter. ***During the term of this research proposal, we will start by identifying the best functional probes of the SLC26A9 permeation pathway. We will then use these probes to define the functional characteristics of anion permeation. Furthermore, taking advantage of the high-resolution structure of the prokaryotic SLC26 homologue (SLD26Dg; D. geothermalis), we will begin to identify key amino acid residues that form the permeation pathway and evaluate changes in their accessibility. ***Overall Impact: Revealing the functional and molecular details of the SLC26A9 ion channel mechanism of action will help us understand the key structural changes that makes this transporter's scaffold act as a channel. The success of this proposal will uncover the underlying mechanisms of membrane transport at the blurred boundary between channels and transporters. ******As an electrophysiologist, I have been studying ion channel function and structure for the last 7 years, applying sophisticated techniques and approaches to explore their mechanism of function. The breadth of my research program is thus unique, especially for such a small starting laboratory currently funded only by my Department's Startup funds. It is my hope that the NSERC committee can judge this research program for its potential within the field of ion channel structure and function.

The long-term goal of this project is to develop a detailed molecular understanding of the SLC26A9 anion channel. SLC26A9 belongs to the SLC26 family of membrane transporters which are found in various organisms, from prokaryotes to mammals. The members of the SLC26 family play central roles in the secretion and absorption of nutrients and many anionic substrates such as SO42−, Cl−, HCO3− or I− which are essential for the cell's physiological processes (e.g. swelling and salt homeostasis). As with other members of its family, SLC26A9 has a modular architecture consisting of two halves, each possessing seven transmembrane segments followed by a cytoplasmic sulfate transporter and an antisigma factor antagonist (STAT) domain. At the functional level, studies have proposed that SLC26A9 functions as a Cl− /HCO3− transporter as well as a Cl− channel; therefore, SLC26A9 is thought to have opposite functional properties, mediating active and passive transport, respectively. These discrepancies in the function of SLC26A9 raise several significant questions about its actual functional mode. Our preliminary data demonstrates that SLC26A9 is a Cl− channel and our main goal is to characterise its function through the investigation of its transport mechanism. In this project, we also aim to identify the parts of the protein that form the Cl− transport pathway and the key structural features essential to support its function as a channel rather than a transporter. ***During the term of this research proposal, we will start by identifying the best functional probes of the SLC26A9 permeation pathway. We will then use these probes to define the functional characteristics of anion permeation. Furthermore, taking advantage of the high-resolution structure of the prokaryotic SLC26 homologue (SLD26Dg; D. geothermalis), we will begin to identify key amino acid residues that form the permeation pathway and evaluate changes in their accessibility. ***Overall Impact: Revealing the functional and molecular details of the SLC26A9 ion channel mechanism of action will help us understand the key structural changes that makes this transporter's scaffold act as a channel. The success of this proposal will uncover the underlying mechanisms of membrane transport at the blurred boundary between channels and transporters. ******As an electrophysiologist, I have been studying ion channel function and structure for the last 7 years, applying sophisticated techniques and approaches to explore their mechanism of function. The breadth of my research program is thus unique, especially for such a small starting laboratory currently funded only by my Department's Startup funds. It is my hope that the NSERC committee can judge this research program for its potential within the field of ion channel structure and function.