Our long-term objective is to define the routes and mechanisms of intracellular cholesterol transport. We are studying cholesterol metabolism in cultured mammalian cells and mice using biochemical and genetic techniques.

Niemann-Pick disease type C (NPC)

NPC is an autosomal recessive lysosomal lipid storage disease characterized by progressive central nervous system degeneration. NPC has classically been described as a disease of cholesterol metabolism; however, it is really a complex lipid storage disorder with increases in cholesterol, neutral and acidic glycosphingolipids, sphingomyelin, and phospholipids.

In normal mammalian cells, low-density lipoprotein (LDL) is bound and internalized by cell surface receptors and is hydrolyzed in the endocytic compartment. The cholesterol that is released is transported to the cell surface and endoplasmic reticulum.

In NPC cells, LDL-derived cholesterol accumulates in lysosomes and endosomes, LDL-cholesterol transport from endocytic compartments to other cellular compartments is delayed, and LDL does not elicit normal homeostatic responses. The NPC1 gene has been cloned, but the biological function of the NPC1 protein has not been determined.

Individuals with NPC experience liver disease and neurodegeneration

Most newborns with NPC present with prolonged jaundice and liver enlargement. Some of these infants will die from liver disease within 2 years; in others, the liver dysfunction will resolve. The latter population typically dies in their teen years from brain cell death.

The downstream effects of NPC1 dysfunction that lead to cell death are unknown. The goal of Victoria McGuinness’ doctoral thesis is to identify the signals that lead to liver cell death. This will benefit NPC patients on two levels. First, it will allow us to explore future therapeutic strategies to delay or prevent liver cell death. Second, the information we learn from liver cell death might be translatable to the neuronal cell death suffered by these patients.

NPC1 Protein

95% of NPC patients have mutations in the npc1 gene, which encodes the NPC1 protein. NPC1 is a large, endosomal/lysosomal membrane protein. Loss of NPC1 protein function causes loss of cholesterol trafficking out of endosomes such that free cholesterol accumulates in lysosomes. We have isolated an NPC1 mutant called 4-4-S that has a partial phenotype: free cholesterol does not accumulate in lysosomes but is in fact trafficked elsewhere within the cell. Despite the apparent lack of cholesterol accumulation, these NPC1 mutant cells do have some phenotypes associated with NPC disease: treatment of these cells with LDL does not cause a decrease in the production of enzymes necessary for cholesterol biosynthesis, nor does treatment cause up-regulation of mechanisms that cause cholesterol storage.

We have determined that 4-4-S cells contain NPC1 with an amino acid substitution in the protein’s first lumenal loop. The 4-4-S NPC1 protein is the focus of Naomi Sayre’s doctoral thesis. She hypothesizes that the 4-4-S NPC1 has partial function such that it can enable trafficking of cholesterol out of endosomes/lysosomes, but cannot enable trafficking or signals that moderate cholesterol homeostasis in response to LDL.