PDZK1 Is Required for Maintaining Hepatic Scavenger Receptor, Class B, Type I (SR-BI) Steady State Levels but Not Its Surface Localization or Function

Pontifical Catholic University of Chile, CiudadSantiago, Santiago Metropolitan, Chile
Journal of Biological Chemistry (Impact Factor: 4.57). 09/2006; 281(39):28975-80. DOI: 10.1074/jbc.M603802200
Source: PubMed


PDZK1 is a multi-PDZ domain-containing adaptor protein that binds to the C terminus of the high density lipoprotein receptor, scavenger receptor, class B, type I (SR-BI), and controls the posttranscriptional, tissue-specific expression of this lipoprotein receptor. In the absence of PDZK1 (PDZK1(-/-) mice), murine hepatic SR-BI protein levels are very low (<5% of control). As a consequence, abnormal plasma lipoprotein metabolism ( approximately 1.5-1.7-fold increased total plasma cholesterol carried in both normal size and abnormally large high density lipoprotein particles) resembles, but is not as severely defective as, that in SR-BI(-/-) mice. Here we show that the total plasma cholesterol levels and size distribution of lipoproteins are virtually identical in SR-BI(-/-) and SR-BI(-/-)/PDZK1(-/-) mice, indicating that most, if not all of the effects of PDZK1 on lipoprotein metabolism are likely because of the effects of PDZK1 on SR-BI. Hepatic overexpression of wild-type SR-BI in PDZK1(-/-) mice restored near or greater than normal levels of cell surface-expressed, functional SR-BI protein levels in the livers of SR-BI(-/-)/PDZK1(-/-) mice and consequently restored apparently normal lipoprotein metabolism in the absence of PDZK1. Thus, PDZK1 is important for maintaining adequate steady state levels of SR-BI in the liver but is not essential for cell surface expression or function of hepatic SR-BI.

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    • "PDZK1 (Postsynaptic Density Protein (PSD-95)/Drosophila Discs-Large (Dlg)/Tight-Junction Protein (ZO1)) is a 519 amino acid, 63 kDa adapter protein that contains four PDZ protein-protein interaction domains [44]–[46]. The first and third PDZ domain of PDZK1 interact with the last three amino acids, AKL, of SR-BI's C-terminal cytoplasmic tail [45], [47], [48]. Knockdown of PDZK1 or deletion of the last three amino acids of SR-BI's C-terminal tail has been shown to impair HDL-dependent signaling in endothelial cells [40], [41]. "
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    ABSTRACT: HDL carries biologically active lipids such as sphingosine-1-phosphate (S1P) and stimulates a variety of cell signaling pathways in diverse cell types, which may contribute to its ability to protect against atherosclerosis. HDL and sphingosine-1-phosphate receptor agonists, FTY720 and SEW2871 triggered macrophage migration. HDL-, but not FTY720-stimulated migration was inhibited by an antibody against the HDL receptor, SR-BI, and an inhibitor of SR-BI mediated lipid transfer. HDL and FTY720-stimulated migration was also inhibited in macrophages lacking either SR-BI or PDZK1, an adaptor protein that binds to SR-BI's C-terminal cytoplasmic tail. Migration in response to HDL and S1P receptor agonists was inhibited by treatment of macrophages with sphingosine-1-phosphate receptor type 1 (S1PR1) antagonists and by pertussis toxin. S1PR1 activates signaling pathways including PI3K-Akt, PKC, p38 MAPK, ERK1/2 and Rho kinases. Using selective inhibitors or macrophages from gene targeted mice, we demonstrated the involvement of each of these pathways in HDL-dependent macrophage migration. These data suggest that HDL stimulates the migration of macrophages in a manner that requires the activities of the HDL receptor SR-BI as well as S1PR1 activity.
    PLoS ONE 09/2014; 9(9):e106487. DOI:10.1371/journal.pone.0106487 · 3.23 Impact Factor
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    • "HDL was labeled with [3H]cholesteryl oleate ([3H]CE) (Perkin-Elmer no. NET746L) as previously described [55]. COS cells were transfected and plated as described above and on day 3 the cells were washed twice with prewarmed (37°C) DMEM. "
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    ABSTRACT: PDZK1 is a four PDZ-domain containing cytoplasmic protein that binds to a variety of membrane proteins via their C-termini and can influence the abundance, localization and/or function of its target proteins. One of these targets in hepatocytes in vivo is the HDL receptor SR-BI. Normal hepatic expression of SR-BI protein requires PDZK1 - <5% of normal hepatic SR-BI is seen in the livers of PDZK1 knockout mice. Progress has been made in identifying features of PDZK1 required to control hepatic SR-BI in vivo using hepatic expression of wild-type and mutant forms of PDZK1 in wild-type and PDZK1 KO transgenic mice. Such in vivo studies are time consuming and expensive, and cannot readily be used to explore many features of the underlying molecular and cellular mechanisms. Here we have explored the potential to use either primary rodent hepatocytes in culture using 2D collagen gels with newly developed optimized conditions or PDZK1/SR-BI co-transfected cultured cell lines (COS, HEK293) for such studies. SR-BI and PDZK1 protein and mRNA expression levels fell rapidly in primary hepatocyte cultures, indicating this system does not adequately mimic hepatocytes in vivo for analysis of the PDZK1 dependence of SR-BI. Although PDZK1 did alter SR-BI protein expression in the cell lines, its influence was independent of SR-BI's C-terminus, and thus is not likely to occur via the same mechanism as that which occurs in hepatocytes in vivo. Caution must be exercised in using primary hepatocytes or cultured cell lines when studying the mechanism underlying the regulation of hepatic SR-BI by PDZK1. It may be possible to use SR-BI and PDZK1 expression as sensitive markers for the in vivo-like state of hepatocytes to further improve primary hepatocyte cell culture conditions.
    PLoS ONE 07/2013; 8(7):e69725. DOI:10.1371/journal.pone.0069725 · 3.23 Impact Factor
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    • "SR-BI contains two cytosolic regions, one of approximately 10 amino acids at its N-terminus and the other of ∼40 amino acids at its C-terminus [4]. The terminal 3-4 amino acids of the C-terminal cytosolic domain represents a binding site for an adaptor protein called PDZK1 which plays an important role in protecting SR-BI protein from degradation in hepatocytes [21] [22] [23]. The precise sequences that direct SR-BI towards degradation in the absence of PDZK1 binding remain to be identified; however it is presumed that they reside in the C-terminal cytosolic tail of SR-BI. "
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    ABSTRACT: SR-BI is a cell surface HDL receptor that mediates selective uptake of the lipid cargo of HDL, an important process in hepatocytes, driving reverse cholesterol transport from cells in the artery wall. To facilitate examination of factors that modulate SR-BI activity in hepatocytes, we have generated fluorescent protein-tagged versions of SR-BI that allow for facile monitoring of SR-BI protein levels and distribution in transfected cells. We show that deletion of the C-terminal cytosolic tail does not affect the distribution of SR-BI in HepG2 cells, nor is the C-terminal cytosolic tail required for SR-BI-mediated uptake of HDL lipids. We also demonstrate that the phorbol ester, PMA, increased, while protein kinase C inhibitors reduced SR-BI-mediated HDL lipid uptake in HepG2 cells. These data suggest that protein kinase C may modulate selective uptake of HDL lipids including cholesterol in hepatocytes, thereby influencing hepatic HDL cholesterol clearance and reverse cholesterol transport.
    Cholesterol 01/2011; 2011(2090-1283):687939. DOI:10.1155/2011/687939
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