Abstract
The epithelial anion transporter SLC26A9 contributes to airway surface hydration and gastric acid production. Colocalizing with CFTR, SLC26A9 has been proposed as a target for the treatment of cystic fibrosis. To provide molecular details of its transport mechanism, we present cryo-EM structures and a functional characterization of murine Slc26a9. These structures define the general architecture of eukaryotic SLC26 family members and reveal an unusual mode of oligomerization which relies predominantly on the cytosolic STAS domain. Our data illustrates conformational transitions of Slc26a9, supporting a rapid alternate-access mechanism which mediates uncoupled chloride transport with negligible bicarbonate or sulfate permeability. The characterization of structure-guided mutants illuminates the properties of the ion transport path, including a selective anion binding site located in the center of a mobile module within the transmembrane domain. This study thus provides a structural foundation for the understanding of the entire SLC26 family and potentially facilitates their therapeutic exploitation.
Highlights
Solute Carrier 26 family member A9 (SLC26A9) is a membrane-transport protein that in recent years has gained considerable attention due to its intriguing functional properties and promising therapeutic potential (Balazs and Mall, 2018)
This is observed for a construct only lacking the intervening sequence (IVS) (SLC26A9DIVS), but not for a construct solely lacking the CT (SLC26A9DCT), suggesting that the IVS might be responsible for the retention of the protein in intracellular membranes (Figure 1—figure supplement 2A)
We observed robust transport activity that can be diminished in a dose-dependent manner by a known inhibitor of Slc26a9, implying that neither the removal of the intrinsically disordered regions (IDRs) from the STAS domain nor the purification procedure has compromised the function of Slc26a9T (Figure 1A, Figure 1—figure supplement 2D)
Summary
Solute Carrier 26 family member A9 (SLC26A9) is a membrane-transport protein that in recent years has gained considerable attention due to its intriguing functional properties and promising therapeutic potential (Balazs and Mall, 2018) It is found abundantly in the epithelia of lung and stomach (Chang et al, 2009b; Lohi et al, 2002; Xu et al, 2005), where it mediates luminal electrogenic chloride transport and contributes to airway clearance and the production of gastric acid. An ion channel known as CFTR provides a route for chloride ions to move out of cells, which helps clear harmful material from the airways Mutations affecting this protein cause the mucus lining the airways to become very sticky, leading to a severe disease known as cystic fibrosis. In combination with electrophysiological experiments, our work demonstrates how Slc26a9 mediates the uncoupled transport of Cl– via a rapid alternate-access mechanism
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