Investigation On The Function Of Type A Scavenger Receptor Cytoplasmic Domain

The class A scavenger receptor (SR-A) is a glycoprotein expressed on the cell surface of macrophages that mediates internalization of chemically modified lipoprotein. It was reported that the receptor internalization required the presence of internalization signal motif and the rate of receptor internalization was governed by the pattern of receptor phosphorylation induced by the ligands. However, the role of the cytoplasmic domain in the receptor-mediated endocytosis is not fully characterized. Here we reported the changes in internalization process of the receptor when the whole cytoplasmic domain sequence (150 base pairs) was truncated. Both the full length and truncated were recombinated into PcDNA3.1/HisB vector and were then transfected to CHO cells separately. After incubated with Dil-acetyl-LDL (Dil-AcLDL), the full length SR-A-transfected cells showed a diffuse distribution of the Dil-Ac-LDL in cytoplasm as well as in cell membrane when monitored under laser confocal microscopy. But in the truncated SR-A-transfected CHO cells, Dil-Ac-LDL mostly distributes at the cell surface only. Measurement of uptake of Dil-Ac-LDL by transfected cells with FACS showed that the bind and uptake of the ligand in full length SR-A was higher than that of truncated receptor(1.3 foldincrease). Treatment with Staurosporine and H7 did not changed Dil-Ac-LDL bound and untaken by truncated receptor, which was different from the full length SR-A that increased obviously. These results indicated that cytoplasmic domain regulate the receptor activity of SR-A, in which the phosphorlation or dephosphorlation of the cytoplasmic domain might play a key role. Additional studies using laser confocal microscopy, flow cytometry, and immunoelectron microscopy demonstrated that the truncated MSR-A was able to distribute in the plasma membrane and bind its lipoprotein ligands but failed to internalize the bound ligands. Similar effects of the truncated MSR-A were observed in monocytic THP-1 cells that express native scavenger receptors. Our results demonstrate that the MSR-A cytoplasmic domain is indispensable in mediating lipid internalization and may provide insights into new strategies for the prevention/treatment of atherosclerosis by molecular modification of MSR-A cytoplasmic domain.