Distribution dynamics and functional importance of NHERF1 in regulation of Mrp-2 trafficking in hepatocytes

Serhan Karvar; Jo Suda; Lixin Zhu; Don C Rockey

doi:10.1152/ajpcell.00011.2014

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Distribution dynamics and functional importance of NHERF1 in regulation of Mrp-2 trafficking in hepatocytes

Journal article

Peer reviewed

Distribution dynamics and functional importance of NHERF1 in regulation of Mrp-2 trafficking in hepatocytes

Serhan Karvar, Jo Suda, Lixin Zhu and Don C Rockey

American Journal of Physiology: Cell Physiology, Vol.307(8), pp.C727-C737

10/15/2014

DOI: 10.1152/ajpcell.00011.2014

PMID: 25163515

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Abstract

Na(+)/H(+) exchanger regulatory factor 1 (NHERF1) is a multifunctional scaffolding protein that interacts with receptors and ion transporters in its PDZ domains and with the ezrin-radixin-moesin (ERM) family of proteins in its COOH terminus. The role of NHERF1 in hepatocyte function remains largely unknown. We examine the distribution and physiological significance of NHERF1 and multidrug resistance-associated protein 2 (Mrp-2) in hepatocytes. A WT radixin binding site mutant (F355R) and NHERF1 PDZ1 and PDZ2 domain adenoviral mutant constructs were tagged with yellow fluorescent protein and expressed in polarized hepatocytes to study localization and function of NHERF1. Cellular distribution of NHERF1 and radixin was visualized by fluorescence microscopy. A 5-chloromethylfluorescein diacetate (CMFDA) assay was used to characterize Mrp-2 function. Similar to Mrp-2, WT NHERF1 and the NHERF1 PDZ2 deletion mutant were localized to the canalicular membrane. In contrast, the radixin binding site mutant (F355R) and the NHERF1 PDZ1 deletion mutant, which interacts poorly with Mrp-2, were rarely associated with the canalicular membrane. Knockdown of NHERF1 led to dramatically impaired CMFDA secretory response. Use of CMFDA showed that the NHERF1 PDZ1 and F355R mutants were devoid of a secretory response, while WT NHERF1-infected cells exhibited increased secretion of glutathione-methylfluorescein. The data indicate that NHERF1 interacts with Mrp-2 via the PDZ1 domain of NHERF1 and, furthermore, that NHERF1 is essential for maintaining the localization and function of Mrp-2.

Animals

ATP Binding Cassette Transporter, Subfamily B - genetics

ATP Binding Cassette Transporter, Subfamily B - metabolism

ATP-Binding Cassette Sub-Family B Member 4

Cell Membrane - metabolism

Cytoskeletal Proteins - metabolism

Gene Expression

Hepatocytes - metabolism

Humans

Membrane Proteins - metabolism

Multidrug Resistance-Associated Protein 2

Mutation, Missense

Phosphoproteins - genetics

Phosphoproteins - metabolism

Protein Binding

Protein Transport

Rats

Sodium-Hydrogen Exchangers - genetics

Sodium-Hydrogen Exchangers - metabolism

Details

Title: Subtitle: Distribution dynamics and functional importance of NHERF1 in regulation of Mrp-2 trafficking in hepatocytes
Creators: Serhan Karvar - Medical University of South Carolina
Jo Suda - University of Southern California
Lixin Zhu - University at Buffalo, State University of New York
Don C Rockey - Medical University of South Carolina
Resource Type: Journal article
Publication Details: American Journal of Physiology: Cell Physiology, Vol.307(8), pp.C727-C737
DOI: 10.1152/ajpcell.00011.2014
PMID: 25163515
ISSN: 0363-6143
eISSN: 1522-1563
Language: English
Date published: 10/15/2014
Academic Unit: Internal Medicine
Record Identifier: 9984695837002771

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