Lipoprotein binding preference of CD36 is altered by filipin treatment

The class B scavenger receptor CD36 binds multiple ligands, including oxidized and native lipoprotein species. CD36 and the related receptor SR-B1 have been localized to caveolae, domains that participate in cell signaling, transcytosis, and regulation of cellular cholesterol homeostasis. Previous work has indicated that the ligand preference of CD36 may depend on the cell type in which it is expressed. To determine if the presence or absence of caveolae is the determining factor for lipoprotein preference, we treated CHO-CD36 and C32 cells with filipin. Filipin treatment rapidly increased the binding capacity of CD36 for the native lipoproteins HDL and LDL, but did not affect the binding capacity of CD36 for oxidized LDL. Filipin treatment affected the distribution of caveolin and CD36 suggesting that the presence caveolae may modulate the ligand preference of CD36. However, its molecular mechanism how CD36 and caveolin interaction in regulating lipoprotein transport remains to be further studied

Smyd1b_tv1, a Key Regulator of Sarcomere Assembly, Is Localized on the M-Line of Skeletal Muscle Fibers

12 páginas, 7 figurasBackground Smyd1b is a member of the Smyd family that plays a key role in sarcomere assembly during myofibrillogenesis. Smyd1b encodes two alternatively spliced isoforms, smyd1b_tv1 and smyd1b_tv2, that are expressed in skeletal and cardiac muscles and play a vital role in myofibrillogenesis in skeletal muscles of zebrafish embryos. Methodology/Principal Findings To better understand Smyd1b function in myofibrillogenesis, we analyzed the subcellular localization of Smyd1b_tv1 and Smyd1b_tv2 in transgenic zebrafish expressing a myc-tagged Smyd1b_tv1 or Smyd1b_tv2. The results showed a dynamic change of their subcellular localization during muscle cell differentiation. Smyd1b_tv1 and Smyd1b_tv2 were primarily localized in the cytosol of myoblasts and myotubes at early stage zebrafish embryos. However, in mature myofibers, Smyd1b_tv1, and to a small degree of Smyd1b_tv2, exhibited a sarcomeric localization. Double staining with sarcomeric markers revealed that Smyd1b_tv1was localized on the M-lines. The sarcomeric localization was confirmed in zebrafish embryos expressing the Smyd1b_tv1-GFP or Smyd1b_tv2-GFP fusion proteins. Compared with Smyd1b_tv1, Smyd1b_tv2, however, showed a weak sarcomeric localization. Smyd1b_tv1 differs from Smyd1b_tv2 by a 13 amino acid insertion encoded by exon 5, suggesting that some residues within the 13 aa insertion may be critical for the strong sarcomeric localization of Smyd1b_tv1. Sequence comparison with Smyd1b_tv1 orthologs from other vertebrates revealed several highly conserved residues (Phe223, His224 and Gln226) and two potential phosphorylation sites (Thr221 and Ser225) within the 13 aa insertion. To determine whether these residues are involved in the increased sarcomeric localization of Smyd1b_tv1, we mutated these residues into alanine. Substitution of Phe223 or Ser225 with alanine significantly reduced the sarcomeric localization of Smyd1b_tv1. In contrast, other substitutions had no effect. Moreover, replacing Ser225 with threonine (S225T) retained the strong sarcomeric localization of Smyd1b_tv1. Conclusion/Significance Together, these data indicate that Phe223 and Ser225 are required for the M-line localization of Smyd1b_tv1.This research was supported by research grant No IS-8713-08 from the Israel Binational Agricultural Research and Development Fund, the United States (BARD), and an intercenter collaboration grant (Du-Fang) from University of Maryland Biotechnology Institute. (http://www.bard-isus.com/). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewe

Lipoprotein binding preference of CD36 is altered by filipin treatment-8

Nel A) or mouse serum IgG (Panel B) or unlabeled oxLDL (panel C) and then 10 μg/ml of Dil-labeled OxLDL was added to incubate for a further 2 – 4 hours. The concentrations of anti-CD36 antibody in panel A and of mouse IgG serum in panel B from top to bottom are: 0, 2, 4, 6, 8 and 10 μg/ml respectively. The concentrations of unlabelled OxLDL in panel C from top to bottom are: 20, 40, 100, 200, and 400 μg/ml. Lipoprotein binding to cells (expressed as mean relative fluoresence) was determined by flow cytometry.<p><b>Copyright information:</b></p><p>Taken from "Lipoprotein binding preference of CD36 is altered by filipin treatment"</p><p>http://www.lipidworld.com/content/7/1/23</p><p>Lipids in Health and Disease 2008;7():23-23.</p><p>Published online 26 Jun 2008</p><p>PMCID:PMC2483703.</p><p></p

Lipoprotein binding preference of CD36 is altered by filipin treatment-7

Erslips and incubated with Dil-labeled lipoproteins (as indicated) at 10 ug/ml of lipoprotein in culture medium at 37°C for 2 – 4 hours. Dil-labeled lipoproteins was determined by examining the cell layers using a fluorescence microscope. Typical images were recorded.<p><b>Copyright information:</b></p><p>Taken from "Lipoprotein binding preference of CD36 is altered by filipin treatment"</p><p>http://www.lipidworld.com/content/7/1/23</p><p>Lipids in Health and Disease 2008;7():23-23.</p><p>Published online 26 Jun 2008</p><p>PMCID:PMC2483703.</p><p></p

Lipoprotein binding preference of CD36 is altered by filipin treatment-2

7°C for 30 minutes and then they were exposed to 10 μg/ml of Dil-lipoprotein at 37°C for 4 hours. The cells were fixed with 4% formaldehyde and viewed under fluorescent microscope. Panel B: Cells were treated same as in A and were then assayed for the presence of Dil by flow cytometry. The data in B represents an average result of three independent experiments.<p><b>Copyright information:</b></p><p>Taken from "Lipoprotein binding preference of CD36 is altered by filipin treatment"</p><p>http://www.lipidworld.com/content/7/1/23</p><p>Lipids in Health and Disease 2008;7():23-23.</p><p>Published online 26 Jun 2008</p><p>PMCID:PMC2483703.</p><p></p

Lipoprotein binding preference of CD36 is altered by filipin treatment-6

Ehyde in PBS, and double immunostained with either FITC-conjugated anti-CD36 antibody and rodomin-conjugated anti-caveolin antibody. The stainned cells were examined under confocal microscope. Upper panel: no filipin was treated and lower panel: filinpin was treated showing more CD36 staing in cytoplam. Magnifications 400×.<p><b>Copyright information:</b></p><p>Taken from "Lipoprotein binding preference of CD36 is altered by filipin treatment"</p><p>http://www.lipidworld.com/content/7/1/23</p><p>Lipids in Health and Disease 2008;7():23-23.</p><p>Published online 26 Jun 2008</p><p>PMCID:PMC2483703.</p><p></p

Lipoprotein binding preference of CD36 is altered by filipin treatment-4

D in Materials and Methods. A: Coomassie blue staining of protein profile from purification fractions. Lane 1, whole cell lysate (80 μg); lane 2, post nuclear supernatant (60 μg), lane 3, Optiprep gradient fraction (30 μg), and lane 4, caveolin-enriched membrane fraction (20 μg). The same amount of proteins (30 μg) from each fraction as in Panel A was analyzed with Western blots by anti-CD36 antibody (B) and anti-caveolin antibody(C).<p><b>Copyright information:</b></p><p>Taken from "Lipoprotein binding preference of CD36 is altered by filipin treatment"</p><p>http://www.lipidworld.com/content/7/1/23</p><p>Lipids in Health and Disease 2008;7():23-23.</p><p>Published online 26 Jun 2008</p><p>PMCID:PMC2483703.</p><p></p

A Study on How Methionine Restriction Decreases the Body’s Hepatic and Lipid Deposition in Rice Field Eel (<i>Monopterus albus</i>)

Methionine restriction reduces animal lipid deposition. However, the molecular mechanism underlying how the body reacts to the condition and regulates lipid metabolism remains unknown. In this study, a feeding trial was performed on rice field eel Monopterus albus with six isonitrogenous and isoenergetic feeds that included different levels of methionine (0, 2, 4, 6, 8, and 10 g/kg). Compared with M0 (0 g/kg), the crude lipid and crude protein of M. albus increased markedly in M8 (8 g/kg) (p p p , hsl, mttp, ldlrap, pparα, cpt1, and cpt2 were remarkably downregulated in M8, while srebf2, lpl, moat2, dgat2, hdlbp, srebf1, fas, fads2, me1, pfae, and icdh were markedly upregulated in M8. Moreover, hepatic SREBP1 and FAS protein expression were upregulated significantly in M8 (p M. albus, especially for hepatic lipid deposition, and mainly downregulated hepatic fatty acid metabolism. Besides, gcn2 could be activated under methionine restriction