Boosting endoplasmic reticulum folding capacity reduces unfolded protein response activation and intracellular accumulation of human kidney anion exchanger 1 in Saccharomyces cerevisiae

• Published in: Yeast (Chichester, England) Vol. 38; no. 9; pp. 521 - 534
• Main Authors: Li, Xiaobing, Cordat, Emmanuelle, Schmitt, Manfred J., Becker, Björn
• Format: Journal Article
• Language: English
• Published: England Wiley Subscription Services, Inc 01.09.2021
• Subjects: Adenosine triphosphatase; Animals; Anion Exchange Protein 1, Erythrocyte - genetics; Anion Exchange Protein 1, Erythrocyte - metabolism; Bicarbonates; Carbonic anhydrase II; Cell Line; Cell surface; chaperone; Electron microscopy; Endoplasmic reticulum; Endoplasmic Reticulum - metabolism; ER stress; Humans; Intracellular; Kidney - metabolism; kidney anion exchanger 1 (kAE1); Kidneys; Mammalian cells; Microscopy; plasma membrane; Protein folding; Protein transport; Proteins; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Unfolded Protein Response; unfolded protein response (UPR); Yeast; yeast model organism; ER stress; yeast model organism; chaperone; plasma membrane; unfolded protein response (UPR); kidney anion exchanger 1 (kAE1)
• ISSN: 0749-503X; 1097-0061
• DOI: 10.1002/yea.3652

Human kidney anion exchanger 1 (kAE1) facilitates simultaneous efflux of bicarbonate and absorption of chloride at the basolateral membrane of α‐intercalated cells. In these cells, kAE1 contributes to systemic acid–base balance along with the proton pump v‐H+‐ATPase and the cytosolic carbonic anhydrase II. Recent electron microscopy analyses in yeast demonstrate that heterologous expression of several kAE1 variants causes a massive accumulation of the anion transporter in intracellular membrane structures. Here, we examined the origin of these kAE1 aggregations in more detail. Using various biochemical techniques and advanced light and electron microscopy, we showed that accumulation of kAE1 mainly occurs in endoplasmic reticulum (ER) membranes which eventually leads to strong unfolded protein response (UPR) activation and severe growth defect in kAE1 expressing yeast cells. Furthermore, our data indicate that UPR activation is dose dependent and uncoupled from the bicarbonate transport activity. By using truncated kAE1 variants, we identified the C‐terminal region of kAE1 as crucial factor for the increased ER stress level. Finally, a redistribution of ER‐localized kAE1 to the cell periphery was achieved by boosting the ER folding capacity. Our findings not only demonstrate a promising strategy for preventing intracellular kAE1 accumulation and improving kAE1 plasma membrane targeting but also highlight the versatility of yeast as model to investigate kAE1‐related research questions including the analysis of structural features, protein degradation and trafficking. Furthermore, our approach might be a promising strategy for future analyses to further optimize the cell surface targeting of other disease‐related PM proteins, not only in yeast but also in mammalian cells. Schematic overview of the experimental setup used in the manuscript. Endoplasmic reticulum (ER) chaperone coexpression prevents the intracellular ER accumulation and unfolded protein response (UPR) activation induced by recombinant expression of the human kidney anion exchanger 1 (kAE1). Under the improved ER folding capacity, kAE1 is properly transported to the yeast cell surface. With this optimized model system, high‐throughput screenings of kAE1 trafficking seem to be feasible in the future. Take Away We analysed the intracellular transport of human kAE1 to the yeast plasma membrane. We studied the effect of human kAE1 expression on yeast growth and UPR activation. We investigated the impact of different kAE1 truncation variants on UPR induction We implemented intervention strategies to improve PM targeting of kAE1.