Regulation of the SLC23A3 anion exchanger activity by the NHERF4 PDZ protein

Abstract

[한글]

[영문]The solute linked carrier (SLC) 26A3, also known as the downregulated in adenomas (DRA), functions as a Cl-/HCO3- exchanger and is expressed at the apical membrane of secretory epithelial cells in the intestines, pancreas and salivary glands. In the exploratory yeast two-hybrid assay, the sodium/proton exchanger regulatory factor (NHERF) 4, a PDZ-containing scaffold protein also known as the intestinal and kidney-enriched PDZ protein (IKEPP), was found to interact with the SLC26A3. In this study, the functional role of interaction between NHERF4 and SLC26A3 was investigated using an integrated molecular physiological approach. Immunoprecipitation with the C-terminus-deleted SLC26A3 mutant revealed that the C-terminal PDZ binding motif of SLC26A3 was required for the SLC26A3-NHERF4 interaction. In addition, a direct protein-protein interaction between the C-terminus of SLC26A3 and the third PDZ domain of NHERF4 (NHERF4-PDZ3) was observed in the GST-based pull-down assay. Of note, co-expression of NHERF4 decreased the surface expression of SLC26A3 by accelerating endocytosis and consequently reduced the SLC26A3-mediated Cl-/HCO3- exchange activities. In contrast, knockdown of the NHERF4 expression by treatment with small interfering RNAs increased the DIDS-insensitive Cl-/HCO3- exchange activities in the HT-29 human colonic epithelial cells. Interestingly, modulation of phosphorylation at the NHERF4-PDZ3 altered the intensity of interaction between NHERF4 and SLC26A3. These results imply that NHERF4 is a physiological regulator of SLC26A3 by affecting its surface expression and that phosphorylation of NHERF4-PDZ3 may be one of the important regulatory factors to control the SLC26A3-NHERF4 interaction.