Localization of the PE methylation pathway and SR-BI to the canalicular membrane

Abstract

To better understand the regulation of biliary phospholipid and cholesterol excretion, canalicular membranes were isolated from the livers of C57BL/6J mice and abundant proteins separated by SDS-PAGE and identified by matrix-assisted laser desorption/ionization mass spectrometry. A prominent protein revealed by this analysis was betaine homocysteine methyltransferase (BHMT). This enzyme catalyzes the first step in a three-enzyme pathway that promotes the methylation of phosphatidylethanolamine (PE) to phosphatidylcholine (PC). Immunoblotting confirmed the presence of BHMT on the canalicular membrane, failed to reveal the presence of the second enzyme in this pathway, methionine adenosyltransferase, and localized the third enzyme of the pathway, PE N-methyltransferase (PEMT). Furthermore, immunfluorescence microscopy unambiguously confirmed the localization of PEMT to the canalicular membrane. These findings indicate that a local mechanism exists in or around hepatocyte canalicular membranes to promote phosphatidylethnolamine methylation and PC biosynthesis. Finally, immunoblotting revealed the presence and immunofluorescence microscopy unambiguously localized the scavenger receptor class B type I (SR-BI) to the canalicular membrane. Therefore, SR-BI, which is known to play a role in cholesterol uptake at the hepatocyte basolateral membrane, may also be involved in biliary cholesterol excretion.Based on these findings, a model is proposed in which local canalicular membrane PC biosynthesis in concert with the phospholipid transporter mdr2 and SR-BI, promotes the excretion of phospholipid and cholesterol into the bile.

CTP:Phosphocholine Cytidylyltransferase: Paving the Way from Gene to Membrane

“…These data are consistent with the view that PC from the CDP-choline pathway also contributes to lipoprotein secretion (18,116). The existence of a separate PC pool governed by the PEMT is suggested by the localization of PEMT to the canalicular membrane where it may promote biliary excretion of PC (117). The data indicate that CCT␣ and PEMT have overlapping roles and suggest that it is a quantitative difference in the capacity for PC production that has an impact on liver function.…”

Hepatic Overexpression of Murine Abcb11 Increases Hepatobiliary Lipid Secretion and Reduces Hepatic Steatosis

“…We did not detect any alterations of the biliary cholesterol transporters Abcg5 and Abcg8, despite an increased biliary cholesterol secretion. Similarly, hepatic expressions of the phosphatidylcholine "flippase" Abcb4 and the scavenger receptor SR-BI (Scarb1), which is another membrane protein reported to be localized in the canalicular membrane and to promote the excretion of phospholipid and cholesterol into the bile (38,39), were also unchanged. These data suggest that the increased cholesterol and phosphatidylcholine secretion into bile is coupled quantitatively to bile salt secretion in our model but is not due to transcriptional regulation of FIG.…”

SR-BI Undergoes Cholesterol-stimulated Transcytosis to the Bile Canaliculus in Polarized WIF-B Cells

“…Second, Silver et al (8) and Wustner et al (31) have proposed that selective depletion of cholesterol destined for transcytotic delivery occurs as HDL undergoes recycling in hepatocytes. Third, Sehayek et al (13) suggested that BC-localized SR-BI mediates cholesterol efflux to phospholipid micelles.…”

“…SR-BI also localizes on the apical domain in gall bladder epithelial cells (11), testicular Sertoli cells (12), isolated primary mouse hepatocytes (13), primary mouse hepatocyte couplets (8), and hepatic tissues sections (14) and has been shown to undergo regulated transcytotic movement in polarized Madin-Darby canine kidney cells (15).…”