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Down regulation of small intestinal ion transport in PDZK1- (CAP70/NHERF3) deficient mice
Jutta Hillesheim1, Brigitte Riederer, Biguang Tuo
1Department of Gastroenterology, Hepatology, and Endocrinology, Hannover Medical School, Carl-Neuberg-Strasse 1, 30625 Hannover, Germany.
Insights
The PDZ-binding protein PDZK1 regulates intestinal electrolyte transport. Its absence in mice impairs sodium absorption and mildly reduces CFTR function, indicating a crucial role in salt balance.
Area of Science:
- Physiology
- Molecular Biology
- Gastroenterology
Background:
- PDZ-binding protein PDZK1 interacts with key intestinal anion transporters like CFTR and NHE3.
- These transporters are vital for intestinal anion secretion and salt absorption.
- The role of PDZK1 in regulating these processes in the native small intestine was previously unclear.
Purpose of the Study:
- To investigate the function of PDZK1 in regulating electrolyte transport in the murine small intestine.
- To determine the impact of PDZK1 deficiency on sodium and bicarbonate transport.
- To assess the expression and localization of relevant transport proteins in the absence of PDZK1.
Main Methods:
- Isolated murine small intestine segments were used to measure short circuit current (I(SC)) and bicarbonate secretion (J(HCO-)(3)).
- Radioactive sodium (22Na+) flux assays assessed sodium absorption.
- Quantitative PCR and Western blotting analyzed mRNA and protein levels of NHE3, CFTR, and PDZK1; immunohistochemistry determined NHE3 localization.
Main Results:
- PDZK1 deficiency in mice significantly reduced basal and forskolin-inhibited sodium absorption in the jejunum.
- A mild reduction in forskolin-stimulated bicarbonate secretion and I(SC) was observed in the duodenum of PDZK1-deficient mice.
- NHE3 mRNA levels were elevated in PDZK1-deficient mice, suggesting altered protein turnover rather than abundance.
Conclusions:
- PDZK1 plays a significant role in regulating electroneutral sodium absorption in the murine small intestine.
- While PDZK1 deficiency causes a mild defect in maximal CFTR activation, it severely impacts sodium absorption.
- These findings highlight PDZK1's importance in maintaining intestinal salt and fluid homeostasis.
Abstract:
The PDZ-binding protein PDZK1 (CAP70/PDZ-dc-1/NHERF3) in vitro binds to cystic fibrosis transmembrane conductance regulator (CFTR), the anion exchangers SLC26A3 and SLC26A6 and the Na(+)/H(+) exchanger NHE3, all of which are major transport proteins for intestinal anion secretion and salt absorption. This study was undertaken to search for a role of PDZK1 in regulating electrolyte transport in native murine small intestine. Short circuit current (I (SC)) and HCO-(3) secretory rate (J(HCO-)(3)) were measured to assess electrogenic anion secretion; (22)Na(+) fluxes to assess sodium absorption in isolated small intestine. NHE3, CFTR, as well as NHERF1, NHERF2, and PDZK1 messenger RNA (mRNA) expression levels, and NHE3 total enterocyte and brush border membrane (BBM) protein abundance were determined by quantitative polymerase chain reaction (PCR) and Western analysis. NHE3 localization was performed by immunohistochemistry. In pdzk1 -/- jejunal mucosa, basal net Na(+) absorption as well as the inhibition of Na(+) absorption by forskolin was significantly reduced. In pdzk1 -/- duodenal mucosa, identical basal I (SC) and (J(HCO-)(3)) but a significant, yet mild, reduction of forskolin-stimulated Delta(J(HCO-)(3)) and DeltaI (SC) was observed compared to +/+ tissue. Tissue conductance, morphological features, and the DeltaI (SC) and increase in (22)Na(+) absorption in response to luminal glucose was identical in pdzk1 +/+ and -/- small intestine, ruling out a general absorptive defect. While CFTR mRNA expression levels were unchanged, NHE3 mRNA expression levels were significantly increased in small intestinal mucosa of pdzk1 -/- mice. Total enterocyte and BBM abundance was not significantly different, suggesting an increased NHE3 turnover, possibly due to reduced NHE3 membrane retention time. Lack of the PDZ-adapter protein PDZK1 in murine small intestine causes a mild reduction in maximal CFTR activation, but a severe defect in electroneutral Na(+) absorption.
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