N-glycosylation and topology of the human SLC26 family of anion transport membrane proteins.

Jing Li, Fan Xia, Reinhart A F Reithmeier

American journal of physiology. Cell physiology 2014 May 15

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The human solute carrier (SLC26) family of anion transporters consists of 10 members (SLCA1-11, SLCA10 being a pseudogene) that encode membrane proteins containing ~12 transmembrane (TM) segments with putative N-glycosylation sites (-NXS/T-) in extracellular loops and a COOH-terminal cytosolic STAS domain. All 10 members of the human SLC26 family, FLAG-tagged at the NH2 terminus, were transiently expressed in HEK-293 cells. While most proteins were observed to contain both high-mannose and complex oligosaccharides, SLC26A2 was mainly in the complex form, SLC26A4 in the high-mannose form, and SLC26A8 was not N-glycosylated. Mutation of the putative N-glycosylation sites showed that most members contain multiple N-glycosylation sites in the second extracytosolic (EC) loop, except SLC26A11, which was N-glycosylated in EC loop 4. Immunofluorescence staining of permeabilized cells localized the proteins to the plasma membrane and the endoplasmic reticulum, with SLC26A2 highly localized to the plasma membrane. N-glycosylation was not a necessary requirement for cell surface expression as the localization of nonglycosylated proteins was similar to their wild-type counterparts, although a lower level of cell-surface biotinylation was observed. No immunostaining of intact cells was observed for any SLC26 members, demonstrating that the NH2-terminal FLAG tag was located in the cytosol. Topological models of the SLC26 proteins that contain an even number of transmembrane segments with both the NH2 and COOH termini located in the cytosol and utilized N-glycosylation sites defining the positions of two EC loops are presented. Copyright © 2014 the American Physiological Society.

Citation

Jing Li, Fan Xia, Reinhart A F Reithmeier. N-glycosylation and topology of the human SLC26 family of anion transport membrane proteins. American journal of physiology. Cell physiology. 2014 May 15;306(10):C943-60