Browsing by Author "Ananthanarayanan, Meenakshisundaram"

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    Localization of the Sodium-Taurocholate cotransporting polypeptide in membrane rafts and modulation of its activity by cholesterol in vitro
    (ELSEVIER SCIENCE BV, 2008) Molina, Hector; Azocar, Lorena; Ananthanarayanan, Meenakshisundaram; Arrese, Marco; Miquel, Juan Francisco
    Background: The relevance of discrete localization of hepatobiliary transporters in specific membrane microdomains is not well known. Aim: To determine whether the Na+/taurocholate cotransporting polypeptide (Ntcp), the main hepatic sinusoidal bile salt transporter, is localized in specific membrane microdomains. Methods: Presence of Ntcp in membrane rafts obtained from mouse liver was studied by immunoblotting and immunofluorescence. HEK-293 cells stably transfected with rat Ntcp were used for in vitro studies. Expression, localization and function of Ntcp in these cells were assessed by immunoblotting, immunofluorescence and biotinylation studies and Na+-dependent taurocholate uptake assays, respectively. The effect of cholesterol depletion/repletion assays on Ntcp function was also investigated. Results: Ntcp localized primarily to membrane rafts in in vivo studies and localized partially in membrane rafts in transfected HEK-293 cells. In these cells, membrane cholesterol depletion resulted in a shift of Ntcp localization into non-membrane rafts, which correlated with a 2.5-fold increase in taurocholate transport. Cholesterol repletion shifted back part of Ntcp into membrane rafts, and normalized taurocholate transport to values similar to control cells. Conclusion: Ntcp localizes in membrane rafts and its localization and function are regulated by membrane cholesterol content. This may serve as a novel regulatory mechanism of bile salt transport in liver. (C) 2008 Elsevier B.V. All rights reserved.
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    βklotho: A new kid on the bile acid biosynthesis block
    (2006) Arrese, Marco; Miquel P., Juan Francisco; Ananthanarayanan, Meenakshisundaram
    We have generated a line of mutant mouse that lacks βKlotho, a protein that structurally resembles Klotho. The synthesis and excretion of bile acids were found to be dramatically elevated in these mutants, and the expression of 2 key bile acid synthase genes, cholesterol 7α-hydroxylase (Cyp7a1) and sterol 12α-hydroxylase (Cyp8b1), was strongly upregulated. Nuclear receptor pathways and the enterohepatic circulation, which regulates bile acid synthesis, seemed to be largely intact; however, bile acid–dependent induction of the small heterodimer partner (SHP) NR0B2, a common negative regulator of Cyp7a1 and Cyp8b1, was significantly attenuated. The expression of Cyp7a1 and Cyp8b1 is known to be repressed by dietary bile acids via both SHP-dependent and -independent regulations. Interestingly, the suppression of Cyp7a1 expression by dietary bile acids was impaired, whereas that of Cyp8b1 expression was not substantially altered in βklotho−/− mice. Therefore, βKlotho may stand as a novel contributor to Cyp7a1-selective regulation. Additionally, βKlotho-knockout mice exhibit resistance to gallstone formation, which suggests the potential future clinical relevance of the βKlotho system.