on March 30, 2006
Published online before print March 30, 2006, doi: 10.1161/01.ATV.0000220171.50282.0c
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Submitted on February 11, 2006
Accepted on March 16, 2006
The Di-Leucine Motif Contributes to Class A Scavenger Receptor-Mediated Internalization of Acetylated Lipoproteins
Yaoyu Chen ;
From the Institute of Reproductive Medicine (Y.Y.C., X.H.W., Q.C.), Nanjing Medical University; Atherosclerosis Research Center, Key Laboratory of Human Functional Genomics (Y.Y.C., X.H.W., J.J.B., S.Y., H.B., X.X.G., X.M.B., L.J., Y.J., L.M.F., Q.C.), Nanjing Medical University, Nanjing, People’s Republic of China.
* To whom correspondence should be addressed. E-mail: qichen{at}njmu.edu.cn.
Objective--The di-leucine motif exists in the intracellular domains of certain cell surface receptors, participating in the receptor-mediated endocytosis. The present study was aimed at determining the role of the di-leucine motif in class A scavenger receptor (SR-A)-mediated ligand endocytosis.
Methods and Results--cDNA coding for a mutant (SR-A mutant N3132LM) with deletion of the di-leucine structure was transfected into Chinese hamster ovary (CHO) cells. Compared with wild-type SR-A-expressing cells, the cells expressing the SR-A mutant N3132LM showed a significant decrease in uptake but almost no change in binding of the SR-A ligand acetylated low-density lipoprotein (AcLDL). Western blot analysis revealed coimmunoprecipitation of SR-A mutant and clathrin from the lysates of the mutant but not wild-type CHO cells, suggesting that AcLDL-bound SR-A mutant N3132LM is associated with the clathrin-coated pit of cellular membrane. Removal of the first 27 amino acid residues from the SR-A N-terminus further reduced AcLDL uptake by the cells with the di-leucine motif mutation.
Conclusions--The di-leucine motif of SR-A intracellular domain contributes to the SR-A-mediated cellular internalization of AcLDL. Di-leucine pair exists in the cytoplasmic domain of class A scavenger receptor. The cells expressing di-leucine mutants showed decreased uptake and unchanged binding of AcLDL. The di-leucine pair was not associated to coated pits. It suggests that di-leucine motif acts as a signal sequence to mediate SR-A into cell.
Key words: atherosclerosis • class A scavenger receptor • di-leucine motif • internalization • clathrin-coated pit