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Lipoprotein binding preference of CD36 is altered by filipin treatment.

Zhang J, Chu W, Crandall I - Lipids Health Dis (2008)

Bottom Line: 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.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Bioengineering and Environmental Science, Changsha University, Changsha, Hunan, PR China. jzhang@ccsu.cn

ABSTRACT
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.

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Semi-quantitation of caveolin and CD36 with and without filipin treatment. Caveolin-enriched membrane protein fractions same as in Fig. 5 were analyzed with both anti-CD36 antibody (upper panel in A) and anti-caveolin antibody (lower panel in A) for the two protein presence. B. Quantitative comparison of caveolin and CD36 proteins in final caveolin-enriched membrane fraction from either filipin treated or untreated cells with dot blots. From left to right, 5, 10, 20 and 40 μg/dot of total proteins were loaded.
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Figure 6: Semi-quantitation of caveolin and CD36 with and without filipin treatment. Caveolin-enriched membrane protein fractions same as in Fig. 5 were analyzed with both anti-CD36 antibody (upper panel in A) and anti-caveolin antibody (lower panel in A) for the two protein presence. B. Quantitative comparison of caveolin and CD36 proteins in final caveolin-enriched membrane fraction from either filipin treated or untreated cells with dot blots. From left to right, 5, 10, 20 and 40 μg/dot of total proteins were loaded.

Mentions: To confirm the hypothesis that filipin treatment causes CD36 to dissociate from caveolae-enriched membrane in CHO CD36, we partially purified the caveolin-enriched membrane fraction from CHO-CD36 cell lysate using a detergent-free method [47]. Immunoblotting of the fractions obtained from untreated CHO-CD36 cells with both an anti-CD36 antibody and an anti-caveolin antibody indicated that CD36 was present in the caveolin-enriched membrane faction (Fig. 5). In contrast, when cells were treated with filipin the amount of CD36 in the final two fractionation steps was greatly diminished (lane 3, 4, Fig. 6A). A semi-quantitative comparison of the caveolin-rich membrane fractions from filipin-treated and untreated CHO-CD36 cells indicated that the amount of CD36 in the final purification fractions of filipin treated cells was decreased by about 30 – 40% relative to the untreated samples (Fig 6B). No difference was seen for the amount of caveolin protein present in the filipin-treated and untreated fractions (Fig. 6B) suggesting that the redistribution of CD36 and its change in ligand preference are related events. Double immunoflorescent staining of CHO-CD36 cells with both anti-CD36 and anti-caveolin antibodies showed that cells without filipin treatment, both CD36 and caveolin concentrated around membranes (Fig. 7,-Filipin), and however, when treated with filipin, more CD36 proteins appeared in the cytoplasm (Fig. 7, +Filipin).


Lipoprotein binding preference of CD36 is altered by filipin treatment.

Zhang J, Chu W, Crandall I - Lipids Health Dis (2008)

Semi-quantitation of caveolin and CD36 with and without filipin treatment. Caveolin-enriched membrane protein fractions same as in Fig. 5 were analyzed with both anti-CD36 antibody (upper panel in A) and anti-caveolin antibody (lower panel in A) for the two protein presence. B. Quantitative comparison of caveolin and CD36 proteins in final caveolin-enriched membrane fraction from either filipin treated or untreated cells with dot blots. From left to right, 5, 10, 20 and 40 μg/dot of total proteins were loaded.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2483703&req=5

Figure 6: Semi-quantitation of caveolin and CD36 with and without filipin treatment. Caveolin-enriched membrane protein fractions same as in Fig. 5 were analyzed with both anti-CD36 antibody (upper panel in A) and anti-caveolin antibody (lower panel in A) for the two protein presence. B. Quantitative comparison of caveolin and CD36 proteins in final caveolin-enriched membrane fraction from either filipin treated or untreated cells with dot blots. From left to right, 5, 10, 20 and 40 μg/dot of total proteins were loaded.
Mentions: To confirm the hypothesis that filipin treatment causes CD36 to dissociate from caveolae-enriched membrane in CHO CD36, we partially purified the caveolin-enriched membrane fraction from CHO-CD36 cell lysate using a detergent-free method [47]. Immunoblotting of the fractions obtained from untreated CHO-CD36 cells with both an anti-CD36 antibody and an anti-caveolin antibody indicated that CD36 was present in the caveolin-enriched membrane faction (Fig. 5). In contrast, when cells were treated with filipin the amount of CD36 in the final two fractionation steps was greatly diminished (lane 3, 4, Fig. 6A). A semi-quantitative comparison of the caveolin-rich membrane fractions from filipin-treated and untreated CHO-CD36 cells indicated that the amount of CD36 in the final purification fractions of filipin treated cells was decreased by about 30 – 40% relative to the untreated samples (Fig 6B). No difference was seen for the amount of caveolin protein present in the filipin-treated and untreated fractions (Fig. 6B) suggesting that the redistribution of CD36 and its change in ligand preference are related events. Double immunoflorescent staining of CHO-CD36 cells with both anti-CD36 and anti-caveolin antibodies showed that cells without filipin treatment, both CD36 and caveolin concentrated around membranes (Fig. 7,-Filipin), and however, when treated with filipin, more CD36 proteins appeared in the cytoplasm (Fig. 7, +Filipin).

Bottom Line: 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.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Bioengineering and Environmental Science, Changsha University, Changsha, Hunan, PR China. jzhang@ccsu.cn

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus