AE1 [anion exchanger 1, also known as SLC4A1 (solute carrier family 4, anion exchanger, member 1) and band 3 (erythrocyte membrane protein band 3)] is a major membrane glycoprotein expressed in human erythrocytes where it mediates the exchange of chloride and bicarbonate across the plasma membrane. Glycophorin A (GPA) is a sialoglycoprotein that associates with AE1 in erythrocytes forming the Wrb (Wright b) blood group antigen. These two integral proteins may also form a complex during biosynthesis, with GPA facilitating the cell surface expression of AE1. This study investigates the interaction of GPA with AE1 in K562 cells, a human erythroleukaemic cell line that expresses GPA, and the role of GPA in the cell surface expression of AE1. In K562 cells, GPA was dimeric and N- and O-glycosylated similar to erythroid GPA. GPA was localized at the cell surface, but also localized to the Golgi. AE1 expressed in K562 cells contained both complex and high-mannose oligosaccharides, and co-localized with GPA at the cell surface and in the endoplasmic reticulum (ER). The Wrb antigen was detected at the cell surface of AE1-transfected K562 cells, indicating the existence of an AE1–GPA complex. Immunofluorescence and co-immunoprecipitation studies using AE1 and an ER-localized hereditary spherocytosis mutant (R760Q AE1) showed that GPA and AE1 could interact in the ER. GPA knockdown by shRNAs (small-hairpin RNAs), however, had no effect on the level of cell surface expression of AE1. The results indicate that AE1 and GPA form a complex in the ER of human K562 cells, but that both proteins can also traffic to the cell surface independently of each other.
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Research Article|
July 15 2009
Interaction of anion exchanger 1 and glycophorin A in human erythroleukaemic K562 cells
Allison J. Pang;
Allison J. Pang
1Department of Biochemistry, University of Toronto, 1 King's College Circle, Medical Sciences Building, Toronto, Canada, M5S 1A8
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Reinhart A. F. Reithmeier
Reinhart A. F. Reithmeier
1
1Department of Biochemistry, University of Toronto, 1 King's College Circle, Medical Sciences Building, Toronto, Canada, M5S 1A8
1To whom correspondence should be addressed (r.reithmeier@utoronto.ca).
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Publisher: Portland Press Ltd
Received:
February 25 2009
Revision Received:
May 06 2009
Accepted:
May 13 2009
Accepted Manuscript online:
May 13 2009
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2009 Biochemical Society
2009
Biochem J (2009) 421 (3): 345–356.
Article history
Received:
February 25 2009
Revision Received:
May 06 2009
Accepted:
May 13 2009
Accepted Manuscript online:
May 13 2009
Citation
Allison J. Pang, Reinhart A. F. Reithmeier; Interaction of anion exchanger 1 and glycophorin A in human erythroleukaemic K562 cells. Biochem J 1 August 2009; 421 (3): 345–356. doi: https://doi.org/10.1042/BJ20090345
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