Ectophosphorylation of CD36 regulates cytoadherence of Plasmodium falciparum to microvascular endothelium under flow conditions.
The adhesion of Plasmodium falciparum-infected erythrocytes (IRBCs) to human dermal microvascular endothelial cells (HDMECs) under flow conditions is regulated by a Src family kinase- and alkaline phosphatase (AP)-dependent mechanism. In this study, we showed that the target of the phosphatase activity is the ectodomain of CD36 at threonine-92 (Thr92). Mouse fibroblasts (NIH 3T3 cells) transfected with wild-type CD36 or a mutant protein in which Thr92 was substituted by Ala supported the rolling and adhesion of IRBCs. However, while the Src family kinase inhibitors PP1 and PP2 and the specific AP inhibitor levamisole significantly reduced IRBC adhesion to wild-type CD36 transfectants as with HDMECs, the inhibitors had no effect on IRBC adhesion to the mutant cells. Using a phosphospecific antibody directed at a 12-amino-acid peptide spanning Thr92, we demonstrated directly that CD36 was constitutively phosphorylated and could be dephosphorylated by exogenous AP. Endothelial CD36 was likewise constitutively phosphorylated. The phosphospecific antibody inhibited IRBC adhesion to HDMECs that could be reversed by preincubating the antibody with the phosphorylated but not the nonphosphorylated peptide. Pretreatment of HDMECs with AP abrogated the effect of PP1 on IRBC adhesion. Collectively, these results are consistent with a critical role for CD36 dephosphorylation through Src family kinase activation in regulating IRBC adhesion to vascular endothelium.
Ho
May
M
Department of Microbiology and Infectious Diseases, University of Calgary, 3330 Hospital Dr. NW, Calgary, Alberta, T2N 4N1 Canada. mho@ucalgary.ca
Hoang
Holly L
HL
Lee
Kristine M
KM
Liu
Naili
N
MacRae
Tara
T
Montes
Laura
L
Flatt
Christine L
CL
Yipp
Bryan G
BG
Berger
Bradley J
BJ
Looareesuwan
Sorrnchai
S
Robbins
Stephen M
SM
D000469:Alkaline Phosphatase / Q000032:analysis / Q000378:metabolism; D000818:Animals; D045163:Antibodies, Phospho-Specific / Q000494:pharmacology; D018955:CD36 Antigens / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D002196:Capillaries / Q000166:cytology; D002448:Cell Adhesion / Q000187:drug effects; D004730:Endothelium, Vascular / Q000276:immunology*; D004912:Erythrocytes / Q000469:parasitology*; D006801:Humans; D007978:Levamisole / Q000494:pharmacology; D051379:Mice; D009154:Mutation; D041681:NIH 3T3 Cells; D010766:Phosphorylation; D010963:Plasmodium falciparum / Q000472:pathogenicity*; D017434:Protein Structure, Tertiary; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D012867:Skin / Q000098:blood supply; D013912:Threonine / Q000235:genetics / Q000378:metabolism; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
C108123:4-amino-5-(4-methylphenyl)-7-(tert-butyl)pyrazolo(3,4-d)pyrimidine; C412373:AG 1879; D045163:Antibodies, Phospho-Specific; D018955:CD36 Antigens; D011720:Pyrazoles; D011743:Pyrimidines; D007978:Levamisole; D013912:Threonine; D019061:src-Family Kinases; D000469:Alkaline Phosphatase