title: Imaging LDL receptor oligomerization during endocytosis using a co-internalization assay.
abstract: Methods to probe receptor oligomerization are useful to understand the molecular mechanisms of receptor signaling. Here we report a fluorescence imaging method to determine receptor oligomerization state in living cells during endocytic internalization. The wild-type receptor is co-expressed with an internalization-defective mutant, and the internalization kinetics of each are independently monitored. If the receptor internalizes as an oligomer, then the wild-type and mutant isoforms will mutually influence each others' trafficking properties, causing co-internalization of the mutant or co-retention of the wild-type at the cell surface. Using this approach, we found that the low density lipoprotein (LDL) receptor internalizes as an oligomer into cells, both in the presence and absence of LDL ligand. The internalization kinetics of the wild-type receptor are not changed by LDL binding. We also found that the oligomerization domain of the LDL receptor is located in its cytoplasmic tail.
authors.forename: Peng
authors.initials: P
authors.lastname: Zou
authors.forename: Alice Y
authors.initials: AY
authors.lastname: Ting
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000032:analysis; D001665:Binding Sites; D001710:Biotin / Q000031:analogs & derivatives / Q000032:analysis / Q000378:metabolism; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D004705:Endocytosis; D005260:Female; D005456:Fluorescent Dyes / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D015870:Gene Expression; D055786:Gene Knockout Techniques; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008077:Lipoproteins, LDL / Q000378:metabolism; D057054:Molecular Imaging / Q000379:methods*; D009154:Mutation; D058105:Polymerization; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs / Q000235:genetics*; D011973:Receptors, LDL / Q000235:genetics / Q000378:metabolism*; D019809:Streptavidin / Q000378:metabolism; D014162:Transfection
chemical_list: D001426:Bacterial Proteins; D005456:Fluorescent Dyes; D008077:Lipoproteins, LDL; D011973:Receptors, LDL; C066114:biotin-streptavidin complex; D001710:Biotin; D019809:Streptavidin