Grp78 Is Involved in Retention of Mutant Low Density Lipoprotein Receptor Protein in the Endoplasmic Reticulum

Malene M Jørgensen,Ole N Jensen,Henrik U Holst,Jens-Jacob Hansen,Thomas J Corydon,Peter Bross,Lars Bolund,Niels Gregersen

doi:10.1074/jbc.m004663200

Abstract

The low density lipoprotein (LDL) receptor is responsible for removing the majority of the LDL cholesterol from the plasma. Mutations in the LDL receptor gene cause the disease familial hypercholesterolemia (FH). Approximately 50% of the mutations in the LDL receptor gene in patients with FH lead to receptor proteins that are retained in the endoplasmic reticulum (ER). Misfolding of mutant LDL receptors is a probable cause of this ER retention, resulting in no functional LDL receptors at the cell surface. However, the specific factors and mechanisms responsible for retention of mutant LDL receptors are unknown. In the present study we show that the molecular chaperone Grp78/BiP co-immunoprecipitates with both the wild type and two different mutant (W556S and C646Y) LDL receptors in lysates obtained from human liver cells overexpressing wild type or mutant LDL receptors. A pulse-chase study shows that the interaction between the wild type LDL receptor and Grp78 is no longer detectable after 2(1/2) h, whereas it persists for more than 4 h with the mutant receptors. Furthermore, about five times more Grp78 is co-immunoprecipitated with the mutant receptors than with the wild type receptor suggesting that Grp78 is involved in retention of mutant LDL receptors in the ER. Overexpression of Grp78 causes no major alterations on the steady state level of active LDL receptors at the cell surface. However, overexpression of Grp78 decreases the processing rate of newly synthesized wild type LDL receptors. This indicates that the Grp78 interaction is a rate-limiting step in the maturation of the wild type LDL receptor and that Grp78 may be an important factor in the quality control of newly synthesized LDL receptors.

Highlights

The low density lipoprotein (LDL) receptor is responsible for removing the majority of the LDL cholesterol from the plasma
50% of the characterized mutations in the LDL receptor gene lead to mutant proteins that are partially or totally retained in the endoplasmic reticulum (ER) [2]
It cannot be excluded that some of these chaperones interact with the LDL receptor, but our results indicate that such interactions are weak if they are present

Summary

Introduction

The low density lipoprotein (LDL) receptor is responsible for removing the majority of the LDL cholesterol from the plasma. Misfolding of mutant LDL receptors is a probable cause of this ER retention, resulting in no functional LDL receptors at the cell surface. Mutations in the LDL receptor gene cause familial hypercholesterolemia (FH), which is an autosomal dominant inherited disorder of lipoprotein metabolism. Missense mutations and small in frame deletions or insertions rarely affect the function of a given protein directly These diseasecausing mutations affect the ability of the proteins to fold into a correct conformation, and they often give rise to premature degradation or aggregation of the mutant proteins. Diseases caused by this kind of molecular pathological mechanism are termed “conformational diseases” [5, 6]. In order to reach a better understanding of the molecular pathogenesis of FH, we embarked on identification of ER quality control components involved in prolonged interaction with mutant, ER-retained, and LDL receptors

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Conclusion