Genetic factors play an important role in contributing to variation in plasma lipid levels across the human population. Large genome-wide association studies (GWAS) in humans have identified more than 100 loci significantly associated with plasma levels of low-density lipoprotein (LDL) cholesterol, high-density lipoprotein (HDL) cholesterol, total cholesterol (TC), and triglycerides. To prioritize genes that are involved in cholesterol metabolism, we developed a systematic approach to leverage genome-wide liver transcriptomic and proteomic data from multiple mouse reference populations along with human lipid GWAS data. We constructed global co-expression networks from twelve distinct mouse liver datasets, encompassing more than 800 unique mice and identified a conserved module of genes highly enriched for cholesterol biosynthesis. Based on replication across datasets and presence in transcript and protein, we prioritized 112 unique genes. Intersection of these 112 prioritized genes with human GWAS data for LDL, HDL, TC, and triglycerides identified 54 genes to be within 100 kilobases of a significant or suggestive significant single nucleotide polymorphism (SNP). Out of the 54 identified genes that overlap with human GWAS data, 29 have well documented biological roles in cholesterol metabolism, such as LDLR , PCSK9 , and INSIG1 . With the 25 identified genes with no described role in cholesterol metabolism, we tested for transcriptional regulation to cholesterol and performed a functional screen by siRNA knockdown. From this analysis, we identified twelve genes that show transcriptional regulation to cholesterol levels and nine genes that are able to modulate cholesterol metabolism when targeted in vitro with siRNA. Five genes out of the 25 prioritized genes show both transcriptional regulation to cholesterol and ability to modulate cholesterol metabolism in vitro. One of these genes, Sestrin1 , we validate in vivo and in vitro as a modulator of cholesterol metabolism. Collectively, through a systematic approach we have identified 25 highly prioritized genes that have no documented role in cholesterol metabolism, five genes are transcriptionally regulated by cholesterol and influence cholesterol metabolism.