Processing of N-linked oligosaccharide depends on its location in the anion exchanger, AE1, membrane glycoprotein

Li, Jing; Quilty, Janne A.; Popov, M.; Reithmeier, Reinhart A.F.

doi:10.1042/bj3490051

Cited by 33 publications

(63 citation statements)

References 19 publications

(29 reference statements)

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“…Similar processing to a complex Nglycan was observed on chicken kAE1 in MDCK cells (AdairKirk et al, 1999). These results contrast with those obtained using human kAE1 transiently transfected into HEK-293 cells, where processing of normal kAE1 and kAE1(R901Stop) did not proceed beyond the core N-glycan (Li et al, 2000;Quilty et al, 2002a;Quilty et al, 2002b). This probably reflects intrinsic differences in trafficking of kAE1 in HEK293 cells and MDCKI cells.…”

Section: Discussioncontrasting

confidence: 55%

“…Cell surface biotinylation Cell surface biotinylation was carried out on metabolically labelled cells as described by Li et al (Li et al, 2000). Cells were treated with non-penetrating biotinylation reagent EZ-link™ Sulfo-NHS-biotin (Pierce, Rockford, IL) at 4°C for 30 minutes, total kAE1 was then immunoprecipitated and the fraction of biotinylated kAE1 determined by binding to streptavidin beads.…”

Section: Cell Transfectionmentioning

confidence: 99%

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Regions of human kidney anion exchanger 1 (kAE1) required for basolateral targeting of kAE1 in polarised kidney cells: mis-targeting explains dominant renal tubular acidosis (dRTA)

Toye

Banting

Tanner

2004

Journal of Cell Science

110

178

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Distal renal tubular acidosis (dRTA) is characterised by defective acid secretion by kidney α-intercalated cells. Some dominantly inherited forms of dRTA result from anion exchanger 1 (AE1) mutations. We have developed a stably transfected cell model for the expression of human kidney AE1 (kAE1) and mutant kAE1 proteins in MDCKI cells. Normal kAE1 was delivered to the plasma membrane of non-polarised cells and to the basolateral membrane of polarised cells. The AE1 N-glycan was processed to a complex form. Surprisingly, expression of kAE1 increased the permeability of the paracellular barrier of polarised MDCKI monolayers. All dominant dRTA mutations examined altered the targeting of kAE1 in MDCKI cells. The mutant proteins kAE1(R589H), kAE1(S613F) and kAE1(R901Stop) were retained in the ER in non-polarised cells, but the kAE1(R901Stop) protein was also present in late endosomes/lysosomes. The complex N-glycan of kAE1(R901Stop) was larger than that of normal kAE1. In polarised cells, the mutant kAE1(R901Stop) was mis-targeted to the apical membrane, while the kAE1(R589H) and kAE1(S613F) mutants did not reach the cell surface. These results demonstrate that dominant dRTA mutations cause aberrant targeting of kAE1 in polarised kidney cells and provide an explanation for the origin of dominant dRTA. Our data also demonstrate that the 11 C-terminal residues of kAE1 contain a tyrosine-dependent basolateral targeting signal that is not recognised by μ1B-containing AP-1 adaptor complexes. In the absence of the N-terminus of kAE1, the C-terminus was not sufficient to localise kAE1 to the basolateral membrane. These results suggest that a determinant within the kAE1 N-terminus co-operates with the C-terminus for kAE1 basolateral localisation.

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Section: Discussioncontrasting

confidence: 55%

Section: Cell Transfectionmentioning

confidence: 99%

Regions of human kidney anion exchanger 1 (kAE1) required for basolateral targeting of kAE1 in polarised kidney cells: mis-targeting explains dominant renal tubular acidosis (dRTA)

Toye

Banting

Tanner

2004

Journal of Cell Science

110

178

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“…This indicates that the PEN-2 S93N mutants is transported to the Golgi complex but fails to become endo H-resistant, because the carbohydrate structure is not modified by Golgi-associated enzymes. In this regard, PEN-2 S93N is similar to certain other glycoproteins, including Nct, which contain one or more Nlinked carbohydrate chains that are not processed by Golgiassociated enzymes even though they transit this organelle (24,35). It may be of interest to introduce carbohydrate addition sites in other regions of the PEN-2 C-terminal domain in the hopes of obtaining a glycosylated version of PEN-2 that is fully processed and so can be used as a marker for intracellular transport events.…”

Section: Discussionmentioning

confidence: 95%

Membrane Topology of γ-Secretase Component PEN-2

Crystal

Morais

Pierson

et al. 2003

Journal of Biological Chemistry

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PEN-2 is an integral membrane protein that is a necessary component of the ␥-secretase complex, which is central in the pathogenesis of Alzheimer's disease and is also required for Notch signaling. In the absence of PEN-2, Notch signaling fails to guide normal development in Caenorhabditis elegans, and amyloid ␤ peptide is not generated from the amyloid precursor protein.Human PEN-2 is a 101-amino acid protein containing two putative transmembrane domains. To understand its interaction with other ␥-secretase components, it is important to know the membrane topology of each member of the complex. To characterize the membrane topology of PEN-2, we introduced single amino acid changes in each of the three hydrophilic regions of PEN-2 to generate N-linked glycosylation sites. We found that the N-linked glycosylation sites present in the N-and C-terminal domains of PEN-2 were utilized, whereas a site in the hydrophilic "loop" region connecting the two transmembrane domains was not. The addition of a carbohydrate structure in the N-terminal domain of PEN-2 prevented association with presenilin 1, whereas glycosylation in the C-terminal region of PEN-2 did not, suggesting that the N-terminal domain is important for interactions with presenilin 1. Immunofluorescence microscopy with selective permeabilization of the plasma membrane of cells expressing epitope-tagged forms of PEN-2 confirmed the lumenal location of both the N and C termini. A protease protection assay also demonstrated that the loop domain of PEN-2 is cytosolic. Thus, PEN-2 spans the membrane twice, with the N and C termini facing the lumen of the endoplasmic reticulum.

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“…Non-glycosylated AE1 can traffic to the plasma membrane in transfected HEK cells (34) and when expressed in Xenopus oocytes (35), suggesting that an oligosaccharide-dependent interaction with CNX or CRT is not essential for cellsurface expression. Our results indicate that although CRT is maintained during the terminal differentiation of red cells, it likely does not substitute for the loss of CNX.…”

Section: Discussionmentioning

confidence: 99%

Loss of Specific Chaperones Involved in Membrane Glycoprotein Biosynthesis during the Maturation of Human Erythroid Progenitor Cells

Patterson

Kang

et al. 2009

Journal of Biological Chemistry

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The production of erythrocytes requires the massive synthesis of red cell-specific proteins including hemoglobin, cytoskeletal proteins, as well as membrane glycoproteins glycophorin A (GPA) and anion exchanger 1 (AE1). We found that during the terminal differentiation of human CD34؉ erythroid progenitor cells in culture, key components of the endoplasmic reticulum (ER) protein translocation (Sec61␣), glycosylation (OST48), and protein folding machinery, chaperones BiP, calreticulin (CRT), and Hsp90 were maintained to allow efficient red cell glycoprotein biosynthesis. Unexpected was the loss of calnexin (CNX), an ER glycoprotein chaperone, and ERp57, a protein-disulfide isomerase, as well as a major decrease of the cytosolic chaperones, Hsc70 and Hsp70, components normally involved in membrane glycoprotein folding and quality control. AE1 can traffic to the cell surface in mouse embryonic fibroblasts completely deficient in CNX or CRT, whereas disruption of the CNX/CRT-glycoprotein interactions in human K562 cells using castanospermine did not affect the cell-surface levels of endogenous GPA or expressed AE1. These results demonstrate that CNX and ERp57 are not required for major glycoprotein biosynthesis during red cell development, in contrast to their role in glycoprotein folding and quality control in other cells.

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Processing of N-linked oligosaccharide depends on its location in the anion exchanger, AE1, membrane glycoprotein

Cited by 33 publications

References 19 publications

Regions of human kidney anion exchanger 1 (kAE1) required for basolateral targeting of kAE1 in polarised kidney cells: mis-targeting explains dominant renal tubular acidosis (dRTA)

Regions of human kidney anion exchanger 1 (kAE1) required for basolateral targeting of kAE1 in polarised kidney cells: mis-targeting explains dominant renal tubular acidosis (dRTA)

Membrane Topology of γ-Secretase Component PEN-2

Loss of Specific Chaperones Involved in Membrane Glycoprotein Biosynthesis during the Maturation of Human Erythroid Progenitor Cells

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