Student Theses and Dissertations

Date of Award

2009

Document Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

RU Laboratory

Breslow Laboratory

Abstract

Regulation of cholesterol levels within cells and organisms is crucial for their survival. Maintenance of cellular cholesterol homeostasis occurs mainly at the transcriptional level, via the Sterol Regulatory Element Binding Proteins (SREBP), the Liver X Receptors (LXR), and the Unfolded Protein Response (UPR)/ER Stress Response. This thesis attempts to characterize the in vivo physiological role of novel downstream target genes of these transcription factors: StARD5 and ADAM11. This thesis outlines the development and characterization of a floxed mouse allele of the UPR-upregulated, reticuloendothelial gene StARD5, as well as two different transgenic mouse lines, one which expresses the human StARD5 gene in the liver and one which expresses the human StARD5 in the reticuloendothelial system. We show that StARD5 plays an indispensable role in early embryogenesis since homozygous knockouts are not viable. Haploinsufficiency in the adult mouse does not alter plasma cholesterol or triglyceride levels, liver cholesterol levels, gallbladder bile composition, glucose tolerance, cholesterol efflux from macrophages, or atherosclerosis susceptibility. Two isoforms of the ADAM11 gene were identified, differing only in their cytoplasmic tails, with one highly expressed in the liver and the other highly expressed in the brain. The liver isoform of ADAM11 was induced by dietary cholesterol and the LXR transcription factor. In order to study the function of this gene, this thesis describes the development of a floxed mouse allele of ADAM11. Utilizing low and high cholesterol and cholic acid containing diets and the ADAM11 knockout, we did not find a phenotype for ADAM11 with respect to plasma cholesterol or triglyceride levels, liver lipid levels, gallbladder bile composition, glucose tolerance, or atherosclerosis susceptibility. We show that loss of ADAM11 in mice fed a high cholesterol diet lowers free plasma campesterol levels, without having an effect on total campesterol levels or other phytosterols. Using a yeast two-hybrid screen, we identified 3HAO as a putative ADAM11 cytoplasmic tail binding partner, which interacted non-specifically with the two cytoplasmic tail forms. ADAM11 knockout mice were previously shown to have a neurological phenotype to which we now add absence-like epilepsy. Finally, this thesis also outlines the development of a floxed mouse allele of the dietary cholesterol-downregulated, SREBP-upregulated liver gene StARD4; initial studies indicate that a full StARD4 knockout is viable.

Comments

A Thesis Presented to the Faculty of The Rockefeller University in Partial Fulfillment of the Requirements for the degree of Doctor of Philosophy

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