Do clustered β-propeller domains within the N-terminus of LRP1 play a functional role?

• Published in: Biochimica et biophysica acta Vol. 1721; no. 1; pp. 139 - 151
• Main Authors: Sun, Fengcheng, Avramoglu, Rita Kohen, Vassiliou, Gerard, Brown, Robert J., Ko, Kerry W.S., McPherson, Ruth, Yao, Zemin
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 18.01.2005
• Subjects: Animals; CHO Cells; CHO LRP1-null cell; COS Cells; Cricetinae; Heat-Shock Proteins - metabolism; Ligands; Low Density Lipoprotein Receptor-Related Protein-1 - chemistry; Low Density Lipoprotein Receptor-Related Protein-1 - physiology; LRP minireceptor; Molecular Chaperones - metabolism; Protein Folding; Protein Transport; Trafficking; β-propeller; RAP; PBS; LRP1; α 2M; β-propeller; EGF; Trafficking; endo H; PAGE; CHO; LRP minireceptor; ER; DMEM; BSA; TCA; FBS; HBSS; CHO LRP1-null cell; LDLR; PNGaseF
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/j.bbagen.2004.10.014

The six β-propellers located within the N-terminus of low density lipoprotein receptor-related protein 1 (LRP1) are arranged in two clusters that contain two and four β-propellers, respectively. Working with LRP1 deletion mutants, we found that randomly removing large segments of amino acid sequences did not affect the intracellular trafficking of LRP1 as long as the clustered β-propeller domains were retained. However, deletion mutants with crippled β-propeller clusters invariably exhibited retarded exit from the endoplasmic reticulum (ER). To determine potential functions of the clustered β-propellers, we generated a series of deletion mutants in which the β-propellers were systematically removed from the C-terminal end of the second cluster. The resulting minireceptors, designated LRPβ1–6, β1–5, β1–4, β1–3, and β1–2 containing decreasing numbers of the β-propellers, were stably expressed in LRP1-null CHO cells. Binding/degradation assays with receptor-associated protein or α 2-macroglobulin showed that removing one or more β-propellers had little effect on binding or degradation of these ligands. However, minireceptors containing odd number of β-propellers (i.e., LRPβ1–3 and β1–5) showed prolonged retention within the ER and remained endoglycosidase H-sensitive, whereas minireceptors containing even number of β-propellers (i.e., LRPβ1–2, β1–4 and β1–6) exited ER at variable rates. Cell surface biotinylation experiments showed that LRPβ1–3 was absent from the cell surface. Prolonged retention of LRPβ1–3 within the ER was accompanied by increased association with molecular chaperone Grp78/Bip. These results suggest that the clustered β-propellers may play a role in folding and intracellular trafficking of LRP1.