BACKGROUND & AIMS: Ezrin-radixin-moesin proteins are cross-linkers between the plasma membrane and actin filaments. Radixin, the dominant ezrin-radixin-moesin protein in hepatocytes, has been reported to selectively tether multidrug-resistance-associated protein 2 to the apical canalicular membrane. However, it remains to be determined if this is its primary function. METHODS: An adenovirus-mediated short interfering RNA (siRNA) was used to down-regulate radixin expression in collagen sandwich-cultured rat hepatocytes and morphologic and functional changes were characterized quantitatively. RESULTS: In control cultures, an extensive bile canalicular network developed with properly localized apical and basolateral transporters that provided for functional excretion of fluorescent cholephiles into the bile canalicular lumina. siRNA-induced suppression of radixin was associated with a marked reduction in the canalicular membrane structure as observed by differential interference contrast microscopy and F-actin staining, in contrast to control cells exposed to adenovirus encoding scrambled siRNA. Indirect immunofluorescence showed that apical transporters (multidrug-resistance-associated protein 2, bile salt export pump, and multidrug-resistance protein 1) dissociated from their normal location at the apical membrane and were found largely associated with Rab11-containing endosomes. Localization of the basolateral membrane transporter, organic anion transporting polypeptide 2 (Oatp2), was not affected. Consistent with this dislocation of apical transporters, the biliary excretion of glutathione-methylfluorescein and cholylglycylamido-fluorescein was decreased significantly in the radixin-deficient cells, but not in the control siRNA cells. CONCLUSIONS: Radixin is essential for maintaining the polarized targeting and/or retaining of canalicular membrane transporters and is a critical determinant of the overall structure and function of the apical membrane of hepatocytes.
BACKGROUND & AIMS: Ezrin-radixin-moesin proteins are cross-linkers between the plasma membrane and actin filaments. Radixin, the dominant ezrin-radixin-moesin protein in hepatocytes, has been reported to selectively tether multidrug-resistance-associated protein 2 to the apical canalicular membrane. However, it remains to be determined if this is its primary function. METHODS: An adenovirus-mediated short interfering RNA (siRNA) was used to down-regulate radixin expression in collagen sandwich-cultured rat hepatocytes and morphologic and functional changes were characterized quantitatively. RESULTS: In control cultures, an extensive bile canalicular network developed with properly localized apical and basolateral transporters that provided for functional excretion of fluorescent cholephiles into the bile canalicular lumina. siRNA-induced suppression of radixin was associated with a marked reduction in the canalicular membrane structure as observed by differential interference contrast microscopy and F-actin staining, in contrast to control cells exposed to adenovirus encoding scrambled siRNA. Indirect immunofluorescence showed that apical transporters (multidrug-resistance-associated protein 2, bile salt export pump, and multidrug-resistance protein 1) dissociated from their normal location at the apical membrane and were found largely associated with Rab11-containing endosomes. Localization of the basolateral membrane transporter, organic anion transporting polypeptide 2 (Oatp2), was not affected. Consistent with this dislocation of apical transporters, the biliary excretion of glutathione-methylfluorescein and cholylglycylamido-fluorescein was decreased significantly in the radixin-deficient cells, but not in the control siRNA cells. CONCLUSIONS:Radixin is essential for maintaining the polarized targeting and/or retaining of canalicular membrane transporters and is a critical determinant of the overall structure and function of the apical membrane of hepatocytes.
Authors: U Beuers; M Bilzer; A Chittattu; G A Kullak-Ublick; D Keppler; G Paumgartner; F Dombrowski Journal: Hepatology Date: 2001-05 Impact factor: 17.425
Authors: Kacper A Wojtal; Mandy Diskar; Friedrich W Herberg; Dick Hoekstra; Sven C D van Ijzendoorn Journal: J Biol Chem Date: 2009-05-22 Impact factor: 5.157
Authors: Raymond Evers; Micheline Piquette-Miller; Joseph W Polli; Frans G M Russel; Jason A Sprowl; Kimio Tohyama; Joseph A Ware; Saskia N de Wildt; Wen Xie; Kim L R Brouwer Journal: Clin Pharmacol Ther Date: 2018-07-12 Impact factor: 6.875