Deletion in Cysteine-Rich Region of LDL Receptor Impedes Transport to Cell Surface in WHHL Rabbit

Abstract

The Watanabe heritable hyperlipidemic (WHHL) rabbit, an animal with familial hypercholesterolemia, produces a mutant receptor for plasma low-density lipoprotein (LDL) that is not transported to the cell surface at a normal rate. Cloning and sequencing of complementary DNA's from normal and WHHL rabbits, shows that this defect arises from an in-frame deletion of 12 nucleotides that eliminates four amino acids from the cysteine-rich ligand binding domain of the LDL receptor. A similar mutation, detected by S1 nuclease mapping of LDL receptor messenger RNA, occurred in a patient with familial hypercholesterolemia whose receptor also fails to be transported to the cell surface. These findings suggest that animal cells may have failsafe mechanisms that prevent the surface expression of improperly folded proteins with unpaired or improperly bonded cysteine residues.Receptors and other cell surface proteins follow a complex path from their sites of synthesis in the rough endoplasmic reticulum to their sites of function in the plasma membrane (1). The orderly progression of such proteins through the endoplasmic reticulum and the Golgi complex was elucidated through study of model proteins, such as those of lipid-enveloped viruses. The signals that direct this movement and the fail-safe mechanisms that prevent denatured proteins from reaching the cell surface are still largely unknown. One way to solve the problem is by studying mutant proteins that do not reach the cell surface. Such mutations have been created artificially through in vitro mutagenesis of genes encoding viral envelope proteins (2). A second approach is through study of naturally occurring mutations in which transport is blocked. In human and animal cells, the most informative group of these mutations occurs in the gene for the low density lipoprotein (LDL) receptor and gives rise to a genetic disease called familial hypercholesterolemia (FH) (3, 4).FH is an autosomal dominant disorder characterized by an elevation of cholesterol in plasma and severe atherosclerosis (3, 4). The disease is caused by defects in the cell surface receptor for LDL, which is a cholesterol transport protein (5). When the LDL receptor is defective, LDL cannot enter cells by receptor-mediated endocytosis and the lipoprotein accumulates in plasma, eventually producing atherosclerosis (3). Some mutations alter the receptor in such a way that it cannot move from the rough endoplasmic reticulum to the Golgi complex (6, 7). Such transport-deficient mutations have been observed frequently in humans with FH (5- WHHL rabbits are homozygous for a mutant allele that produces an LDL receptor that is of normal apparent molecular size but is transported to the cell surface at only one-tenth the normal rate (7). The newly synthesized receptor receives its normal complement of high mannose N-linked sugars and the core N-acetylgalactosamine of the O-linked sugars (5). However, these carbohydrate chains are not processed to their mature form, apparently because the mutant receptor does no...

Defective processing and binding of low‐density lipoprotein receptors in fibroblasts from a familial hypercholesterolaemic subject

“…The poor processing in the Watanabe heritable hyperlipidaemic rabbit and in a similar FH subject results from a small deletion in the binding region of the receptor [21], but this also prevented the bulk of the receptor from reaching the surface. A larger deletion in the same region reduces the binding of LDL, but not that of j-VLDL 1221.…”

Ultrastructural changes in atherosclerotic plaques following the instillation of airborne particulate matter into the lungs of rabbits

“…WHHL rabbits have a spontaneous four amino acid deletion in the cysteine-rich ligand-binding domain of the low-density lipoprotein (LDL) receptor gene -the same gene associated with human familial hypercholesterolemia (54)(55)(56). In both humans and rabbits, this genetic defect produces a drastic reduction of plasma membrane LDL receptors on endothelial cells and leads to the decreased clearance of lipoproteins, resulting in extremely high levels of LDL in the circulating blood (55,57).…”

Folding intermediates are involved in genetic diseases?

“…Class 2 mutations of LDL receptor (50% of all cases) results in a protein which cannot be secreted from the ER [1,22,23]. Another example is the Tay-Sachs disease which is connected with mutations in lysosomal enzymes, which dimeric forms are finally processed in lysosomes.…”