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(Arteriosclerosis, Thrombosis, and Vascular Biology. 2006;26:1317.)
© 2006 American Heart Association, Inc.

Atherosclerosis and Lipoproteins

The Di-Leucine Motif Contributes to Class A Scavenger Receptor-Mediated Internalization of Acetylated Lipoproteins

Yaoyu Chen; Xiaohua Wang; Jingjing Ben; Shen Yue; Hui Bai; Xiaoxiang Guan; Xiaoming Bai; Li Jiang; Yong Ji; Leming Fan; Qi 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.

Correspondence to Qi Chen, Nanjing Medical University, Nanjing, People’s Republic of China. 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.

Di-leucine pair exists in human, bovine, and rabbit cytoplasmic domains 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