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Polarization of membrane associated proteins in the choroid plexus epithelium from normal and slc4a10 knockout mice.

Christensen IB, Gyldenholm T, Damkier HH, Praetorius J - Front Physiol (2013)

Bottom Line: Anion exchanger 2 abundance is increased in slc4a10 knockout and its anchor protein, α-adducin is almost exclusively found near the basolateral domain.E-cadherin expression is unchanged in the slc4a10 knockout, while small decreases in abundance are observed for its probable adaptor proteins, the catenins.Interestingly, the abundance of the tight junction protein claudin-2 is significantly reduced in the slc4a10 knockouts, which may critically affect paracellular transport in this epithelium.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedicine, Faculty of Health, Aarhus University Aarhus, Denmark.

ABSTRACT
The choroid plexus epithelium (CPE) has served as a model-epithelium for cell polarization and transport studies and plays a crucial role for cerebrospinal fluid (CSF) production. The normal luminal membrane expression of Na(+),K(+)-ATPase, aquaporin-1 and Na(+)/H(+) exchanger 1 in the choroid plexus is severely affected by deletion of the slc4a10 gene that encodes the bicarbonate transporting protein Ncbe/NBCn2. The causes for these deviations from normal epithelial polarization and redistribution following specific gene knockout are unknown, but may be significant for basic epithelial cell biology. Therefore, a more comprehensive analysis of cell polarization in the choroid plexus is warranted. We find that the cytoskeleton in the choroid plexus contains αI-, αII-, βI-, and βII-spectrin isoforms along with the anchoring protein ankyrin-3, most of which are mainly localized in the luminal membrane domain. Furthermore, we find α-adducin localized near the plasma membranes globally, but with only faint expression in the luminal membrane domain. In slc4a10 knockout mice, the abundance of β1 Na(+),K(+)-ATPase subunits in the luminal membrane is markedly reduced. Anion exchanger 2 abundance is increased in slc4a10 knockout and its anchor protein, α-adducin is almost exclusively found near the basolateral domain. The αI- and βI-spectrin abundances are also decreased in the slc4a10 knockout, where the basolateral domain expression of αI-spectrin is exchanged for a strictly luminal domain localization. E-cadherin expression is unchanged in the slc4a10 knockout, while small decreases in abundance are observed for its probable adaptor proteins, the catenins. Interestingly, the abundance of the tight junction protein claudin-2 is significantly reduced in the slc4a10 knockouts, which may critically affect paracellular transport in this epithelium. The observations allow the generation of new hypotheses on basic cell biological paradigms that can be tested experimentally in future studies.

No MeSH data available.


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E-cadherin, α- and β-catenin expression in CPE from slc4a10 wt and ko mice. Mouse brain sections were immunostained for E-cadherin and α- and β-catenin, as indicated. (A) Immunohistochemical detection of E-cadherin in slc4a10 wt and ko, as indicated. Bar graphs on the right show the semi-quantitation of the immunofluorescence in slc4a10 wt and ko mouse CPE (n = 4). (B) Immunolabeling for α-catenin in slc4a10 wt and ko, as indicated. Bar graph on the right show the semi-quantitation of the immunofluorescence in slc4a10 wt and ko mouse CPE (wt: n = 5, ko: n = 3). (C) High magnification micrograph of the subcellular α-catenin staining pattern. The histogram on the right is a linescan of the cellular fluorescence intensity from the basal to the luminal domain. The analyzed line is indicated on the two micrographs on the left. (D) Immunolabeling for β-catenin in slc4a10 wt and ko, as indicated. Bar graph on the right shows the semi-quantitation of the immunofluorescence in wt and ko mouse CPE (wt: n = 5, ko: n = 3). Arrows indicate the luminal membrane, while arrowheads indicate the basolateral membrane. *indicates statistical significance. Cell nuclei were visualized by Topro nuclear staining (blue).
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Figure 9: E-cadherin, α- and β-catenin expression in CPE from slc4a10 wt and ko mice. Mouse brain sections were immunostained for E-cadherin and α- and β-catenin, as indicated. (A) Immunohistochemical detection of E-cadherin in slc4a10 wt and ko, as indicated. Bar graphs on the right show the semi-quantitation of the immunofluorescence in slc4a10 wt and ko mouse CPE (n = 4). (B) Immunolabeling for α-catenin in slc4a10 wt and ko, as indicated. Bar graph on the right show the semi-quantitation of the immunofluorescence in slc4a10 wt and ko mouse CPE (wt: n = 5, ko: n = 3). (C) High magnification micrograph of the subcellular α-catenin staining pattern. The histogram on the right is a linescan of the cellular fluorescence intensity from the basal to the luminal domain. The analyzed line is indicated on the two micrographs on the left. (D) Immunolabeling for β-catenin in slc4a10 wt and ko, as indicated. Bar graph on the right shows the semi-quantitation of the immunofluorescence in wt and ko mouse CPE (wt: n = 5, ko: n = 3). Arrows indicate the luminal membrane, while arrowheads indicate the basolateral membrane. *indicates statistical significance. Cell nuclei were visualized by Topro nuclear staining (blue).

Mentions: Major changes in protein abundances of E-cadherin and the catenins were not expected in the slc4a10 ko CPE, as the major changes in protein expression are observed in relation to the luminal plasma membrane. The micrographs in Figure 9A shows similar labeling intensity and cellular distribution of E-cadherin in slc4a10 wt and ko CPE (p = 0.686, n = 4 and 4 for wt and ko). Nevertheless, as seen in Figure 9B, the abundance of α-catenin is significantly decreased in the slc4a10 ko CPE as compared to the wt (Figure 9B Bar graph, p = 0.0357, n = 5 and 3 for wt and ko, respectively). Furthermore, the cellular localization of α-catenin seems to become more abundant in the lateral membrane domain and less in the basolateral labyrinth (Figure 9C). For β-catenin, similar expression patterns and protein abundances were observed in the two genotypes (Figure 9D, p = 0.143, n = 5 and 3 for wt and ko, respectively).


Polarization of membrane associated proteins in the choroid plexus epithelium from normal and slc4a10 knockout mice.

Christensen IB, Gyldenholm T, Damkier HH, Praetorius J - Front Physiol (2013)

E-cadherin, α- and β-catenin expression in CPE from slc4a10 wt and ko mice. Mouse brain sections were immunostained for E-cadherin and α- and β-catenin, as indicated. (A) Immunohistochemical detection of E-cadherin in slc4a10 wt and ko, as indicated. Bar graphs on the right show the semi-quantitation of the immunofluorescence in slc4a10 wt and ko mouse CPE (n = 4). (B) Immunolabeling for α-catenin in slc4a10 wt and ko, as indicated. Bar graph on the right show the semi-quantitation of the immunofluorescence in slc4a10 wt and ko mouse CPE (wt: n = 5, ko: n = 3). (C) High magnification micrograph of the subcellular α-catenin staining pattern. The histogram on the right is a linescan of the cellular fluorescence intensity from the basal to the luminal domain. The analyzed line is indicated on the two micrographs on the left. (D) Immunolabeling for β-catenin in slc4a10 wt and ko, as indicated. Bar graph on the right shows the semi-quantitation of the immunofluorescence in wt and ko mouse CPE (wt: n = 5, ko: n = 3). Arrows indicate the luminal membrane, while arrowheads indicate the basolateral membrane. *indicates statistical significance. Cell nuclei were visualized by Topro nuclear staining (blue).
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Figure 9: E-cadherin, α- and β-catenin expression in CPE from slc4a10 wt and ko mice. Mouse brain sections were immunostained for E-cadherin and α- and β-catenin, as indicated. (A) Immunohistochemical detection of E-cadherin in slc4a10 wt and ko, as indicated. Bar graphs on the right show the semi-quantitation of the immunofluorescence in slc4a10 wt and ko mouse CPE (n = 4). (B) Immunolabeling for α-catenin in slc4a10 wt and ko, as indicated. Bar graph on the right show the semi-quantitation of the immunofluorescence in slc4a10 wt and ko mouse CPE (wt: n = 5, ko: n = 3). (C) High magnification micrograph of the subcellular α-catenin staining pattern. The histogram on the right is a linescan of the cellular fluorescence intensity from the basal to the luminal domain. The analyzed line is indicated on the two micrographs on the left. (D) Immunolabeling for β-catenin in slc4a10 wt and ko, as indicated. Bar graph on the right shows the semi-quantitation of the immunofluorescence in wt and ko mouse CPE (wt: n = 5, ko: n = 3). Arrows indicate the luminal membrane, while arrowheads indicate the basolateral membrane. *indicates statistical significance. Cell nuclei were visualized by Topro nuclear staining (blue).
Mentions: Major changes in protein abundances of E-cadherin and the catenins were not expected in the slc4a10 ko CPE, as the major changes in protein expression are observed in relation to the luminal plasma membrane. The micrographs in Figure 9A shows similar labeling intensity and cellular distribution of E-cadherin in slc4a10 wt and ko CPE (p = 0.686, n = 4 and 4 for wt and ko). Nevertheless, as seen in Figure 9B, the abundance of α-catenin is significantly decreased in the slc4a10 ko CPE as compared to the wt (Figure 9B Bar graph, p = 0.0357, n = 5 and 3 for wt and ko, respectively). Furthermore, the cellular localization of α-catenin seems to become more abundant in the lateral membrane domain and less in the basolateral labyrinth (Figure 9C). For β-catenin, similar expression patterns and protein abundances were observed in the two genotypes (Figure 9D, p = 0.143, n = 5 and 3 for wt and ko, respectively).

Bottom Line: Anion exchanger 2 abundance is increased in slc4a10 knockout and its anchor protein, α-adducin is almost exclusively found near the basolateral domain.E-cadherin expression is unchanged in the slc4a10 knockout, while small decreases in abundance are observed for its probable adaptor proteins, the catenins.Interestingly, the abundance of the tight junction protein claudin-2 is significantly reduced in the slc4a10 knockouts, which may critically affect paracellular transport in this epithelium.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedicine, Faculty of Health, Aarhus University Aarhus, Denmark.

ABSTRACT
The choroid plexus epithelium (CPE) has served as a model-epithelium for cell polarization and transport studies and plays a crucial role for cerebrospinal fluid (CSF) production. The normal luminal membrane expression of Na(+),K(+)-ATPase, aquaporin-1 and Na(+)/H(+) exchanger 1 in the choroid plexus is severely affected by deletion of the slc4a10 gene that encodes the bicarbonate transporting protein Ncbe/NBCn2. The causes for these deviations from normal epithelial polarization and redistribution following specific gene knockout are unknown, but may be significant for basic epithelial cell biology. Therefore, a more comprehensive analysis of cell polarization in the choroid plexus is warranted. We find that the cytoskeleton in the choroid plexus contains αI-, αII-, βI-, and βII-spectrin isoforms along with the anchoring protein ankyrin-3, most of which are mainly localized in the luminal membrane domain. Furthermore, we find α-adducin localized near the plasma membranes globally, but with only faint expression in the luminal membrane domain. In slc4a10 knockout mice, the abundance of β1 Na(+),K(+)-ATPase subunits in the luminal membrane is markedly reduced. Anion exchanger 2 abundance is increased in slc4a10 knockout and its anchor protein, α-adducin is almost exclusively found near the basolateral domain. The αI- and βI-spectrin abundances are also decreased in the slc4a10 knockout, where the basolateral domain expression of αI-spectrin is exchanged for a strictly luminal domain localization. E-cadherin expression is unchanged in the slc4a10 knockout, while small decreases in abundance are observed for its probable adaptor proteins, the catenins. Interestingly, the abundance of the tight junction protein claudin-2 is significantly reduced in the slc4a10 knockouts, which may critically affect paracellular transport in this epithelium. The observations allow the generation of new hypotheses on basic cell biological paradigms that can be tested experimentally in future studies.

No MeSH data available.


Related in: MedlinePlus