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NHERF-1 binds to Mrp2 and regulates hepatic Mrp2 expression and function.
Man Li1, Wei Wang, Carol J Soroka
1Liver Center, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
The Journal of Biological Chemistry
|April 21, 2010
Summary
Na+/H+ exchanger regulatory factor 1 (NHERF-1) binds to multidrug resistance-associated protein 2 (Mrp2), impacting its expression and function. NHERF-1 deficiency reduces Mrp2 protein levels and impairs bile flow, highlighting NHERF-1
Area of Science:
- Hepatobiliary physiology
- Molecular biology
- Membrane transport
Background:
- Multidrug resistance-associated protein 2 (Mrp2) is crucial for hepatocyte canalicular membrane transport, bile formation, and detoxification.
- Na+/H+ exchanger regulatory factor 1 (NHERF-1) is a PDZ protein that links membrane proteins to the actin cytoskeleton.
- The specific role of NHERF-1 in Mrp2 expression and function is not well understood.
Purpose of the Study:
- To investigate the interaction between Mrp2 and NHERF-1.
- To determine the physiological significance of this interaction in Mrp2 expression and function.
Main Methods:
- Co-immunoprecipitation in co-transfected HEK293 cells to assess Mrp2-NHERF-1 interaction.
- Analysis of Mrp2 mRNA and protein levels in NHERF-1 knockout (NHERF-1(-/-)) mouse liver.
- Immunofluorescence microscopy to evaluate Mrp2 localization in NHERF-1(-/-) hepatocytes.
- Measurement of bile flow, glutathione, and glutathione-methylfluorescein (GS-MF) excretion in NHERF-1(-/-) mice.
Main Results:
- Mrp2 co-precipitated with NHERF-1, requiring Mrp2's PDZ-binding motif.
- NHERF-1(-/-) mouse liver showed reduced Mrp2 protein levels (~50% in whole lysates, ~70% in membrane fractions) without changes in Mrp2 mRNA.
- Mrp2 localization remained at the canalicular membrane, but with weaker immunofluorescence.
- Bile flow was reduced by ~70% in NHERF-1(-/-) mice, accompanied by decreased glutathione (~50%) and GS-MF (~60%) excretion.
- Bile acid and bilirubin excretion were unaffected.
Conclusions:
- NHERF-1 directly binds to Mrp2.
- NHERF-1 plays a critical role in maintaining canalicular Mrp2 protein expression via post-transcriptional mechanisms.
- NHERF-1 is essential for Mrp2-mediated glutathione-dependent, bile acid-independent bile flow.
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