Journal article
Phosphorylation dynamics of radixin in hypoxia-induced hepatocyte injury
American journal of physiology: Gastrointestinal and liver physiology, Vol.308(4), pp.G313-G324
02/15/2015
DOI: 10.1152/ajpgi.00369.2014
PMID: 25501552
Abstract
The most prominent ezrin-radixin-moesin protein in hepatocytes is radixin, which is localized primarily at the canalicular microvilli and appears to be important in regulation of cell polarity and in localizing the multidrug resistance-associated protein 2 (Mrp-2) function. Our aim was to investigate how hypoxia affects radixin distribution and Mrp-2 function. We created wild-type and mutant constructs (in adenoviral vectors), which were expressed in WIF-B cells. The cellular distribution of Mrp-2 and radixin was visualized by fluorescence microscopy, and a 5-chloromethylfluorescein diacetate (CMFDA) assay was used to measure Mrp-2 function. Under usual conditions, cells infected with wild-type radixin, nonphosphorylatable radixin-T564A, and radixin-T564D (active phospho-mimicking mutant) were found to be heavily expressed in canalicular membrane compartment vacuoles, typically colocalizing with Mrp-2. In contrast, after hypoxia for 24 h, both endogenous and overexpressed wild-type radixin and the radixin-T564A mutant were found to be translocated to the cytoplasmic space. However, distribution of the radixin-T564D mutant, which mimics constant phosphorylation, was remarkably different, being associated with canalicular membranes even in hypoxic conditions. This dominant-active construct also prevented dissociation of radixin from the plasma membrane. Hypoxia also led to Mrp-2 mislocalization and caused Mrp-2 to be dissociated from radixin; the radixin phospho-mimicking mutant (T564D) abrogated this effect of hypoxia. Finally, hypoxia diminished the secretory response (measured using the CMFDA assay) in WIF-B cells, and the dominant-active construct (radixin-T567D) rescued this phenotype. Taken collectively, these findings suggest that radixin regulates Mrp-2 localization and function in hepatocytes and is important in hypoxic liver injury.
Details
- Title: Subtitle
- Phosphorylation dynamics of radixin in hypoxia-induced hepatocyte injury
- Creators
- Jo Suda - University of Southern CaliforniaDon C Rockey - Medical University of South CarolinaSerhan Karvar - Medical University of South Carolina
- Resource Type
- Journal article
- Publication Details
- American journal of physiology: Gastrointestinal and liver physiology, Vol.308(4), pp.G313-G324
- DOI
- 10.1152/ajpgi.00369.2014
- PMID
- 25501552
- ISSN
- 0193-1857
- eISSN
- 1522-1547
- Language
- English
- Date published
- 02/15/2015
- Academic Unit
- Internal Medicine
- Record Identifier
- 9984695679602771
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