To elucidate whether Sterol O-acyltransferase (Soat) mediates the absorption and transportation of yolk lipids to the developing embryo, zebrafish soat1 and soat2 were cloned and studied. In the adult zebrafish, soat1 was detected ubiquitously while soat2 mRNA was detected specifically in the liver, intestine, brain and testis. Whole mount in situ hybridization demonstrated that both soat1 and soat2 expressed in the yolk syncytial layer, hatching gland and developing cardiovascular as well as digestive systems, suggesting that Soats may play important roles in the lipid trafficking and utilization during embryonic development. The enzymatic activity of zebrafish Soat2 was confirmed by Oil Red O staining in the HEK293 cells overexpressing this gene, and could be quenched by Soat2 inhibitor Pyripyropene A (PPPA). The zebrafish embryos injected with PPPA or morpholino oligo against soat2 in the yolk showed significantly larger yolk when compared with wild-type embryos, especially at 72 hpf, indicating a slower rate of yolk consumption. Our result indicated that zebrafish Soat2 is catalytically active in synthesizing cholesteryl esters and contributes to the yolk cholesterol trafficking during zebrafish embryogenesis.

The primary risk factor for atherosclerotic cardiovascular disease is LDL cholesterol, which can be reduced by increasing cholesterol excretion from the body. Fecal cholesterol excretion can be driven by a hepatobiliary as well as a non-biliary pathway known as transintestinal cholesterol efflux (TICE). We previously showed that chronic knockdown of the hepatic cholesterol esterifying enzyme sterol O-acyltransferase 2 (SOAT2) increased fecal cholesterol loss via TICE. To elucidate the initial events that stimulate TICE, C57Bl/6 mice were fed a high cholesterol diet to induce hepatic cholesterol accumulation and were then treated for 1 or 2 weeks with an antisense oligonucleotide targeting SOAT2. Within 2 weeks of hepatic SOAT2 knockdown (SOAT2HKD), the concentration of cholesteryl ester in the liver was reduced by 70% without a reciprocal increase in hepatic free cholesterol. The rapid mobilization of hepatic cholesterol stores resulted in a ∼ 2-fold increase in fecal neutral sterol loss but no change in biliary cholesterol concentration. Acute SOAT2HKD increased plasma cholesterol carried primarily in lipoproteins enriched in apoB and apoE. Collectively, our data suggest that acutely reducing SOAT2 causes hepatic cholesterol to be swiftly mobilized and packaged onto nascent lipoproteins that feed cholesterol into the TICE pathway for fecal excretion.

In a large scale screen for skin, hair, and nail abnormalities in null mice generated by The Jackson Laboratory's KOMP center, homozygous mutant Far2tm2b(KOMP)Wtsi/2J (hereafter referrred to as Far2-/-) mice were found to develop focal areas of alopecia as they aged. As sebocytes matured in wildtype C57BL/NJ mice they became pale with fine, uniformly sized clear lipid containing vacuoles that were released when sebocytes disintegrated in the duct. By contrast, the Far2-/- null mice had sebocytes that were similar within the gland but become brightly eosinophilic when the cells entered the sebaceous gland duct. As sebocytes disintegrated, their contents did not readily dissipate. Scattered throughout the dermis, and often at the dermal hypodermal fat junction, were dystrophic hair follicles or ruptured follicles with a foreign body granulomatous reaction surrounding free hair shafts (trichogranuloma). The Meibomian and clitoral glands (modified sebaceous glands) of Far2-/- mice showed ducts dilated to various degrees that were associated with mild changes in the sebocytes as seen in the truncal skin. Skin surface lipidomic analysis revealed a lower level of wax esters, cholesterol esters, ceramides, and diacylglycerols compared to wildtype control mice. Similar changes were described in a number of other mouse mutations that affected the sebaceous glands resulting in primary cicatricial alopecia.

During avian embryonic development, endodermal epithelial cells (EECs) absorb yolk through the yolk sac membrane. Sterol O-acyltransferase (SOAT) is important for esterification and yolk lipid utilization during development. Because the major enzyme for yolk sac membrane cholesteryl ester synthesis is SOAT1, we cloned the avian SOAT1 promoter and elucidated the cellular functions of SOAT1. Treatments with either glucagon, isobutylmethylxanthine (IBMX), an adenylate cyclase activator (forskolin), a cAMP analog (dibutyryl-cAMP), or a low glucose concentration all increased SOAT1 mRNA accumulation in EECs from Japanese quail, suggesting that SOAT1 is regulated by nutrients and hormones through a cAMP-dependent pathway. Activity of protein kinase A (PKA) was increased by IBMX, whereas co-treatment with the PKA inhibitor, H89 negated the increase in PKA activity. Cyclic AMP-induced EECs had greater cholesterol esterification than untreated EECs. By promoter deletion and point-mutation, the cAMP-response element (-349 to -341 bp) was identified as critical in mediating transcription of SOAT1. In conclusion, expression of SOAT1 was regulated by a cAMP-dependent pathway and factors that increase PKA will increase SOAT1 to improve the utilization of lipids in the EECs and potentially modify embryonic growth.

We previously identified two closely linked quantitative trait loci (QTL) on distal chromosome 1 contributing to major variations in plasma cholesterol and triglyceride levels in an intercross derived from C57BL/6 (B6) and C3H/HeJ (C3H) apolipoprotein E-deficient (apoE(-/-)) mice. Soat1, encoding sterol o-acyltransferase 1, is a functional candidate gene located underneath the proximal linkage peak. We sequenced the coding region of Soat1 and identified four single nucleotide polymorphisms (SNPs) between B6 and C3H mice. Two of the SNPs resulted in amino-acid substitutions (Ile147Val and His205Tyr). Functional assay revealed an increased enzyme activity of Soat1 in peritoneal macrophages of C3H mice relative to those of B6 mice despite comparable protein expression levels. Allelic variants of Soat1 were associated with variations in plasma cholesterol and triglyceride levels in an intercross between B6.apoE(-/-) and C3H.apoE(-/-) mice. Inheritance of the C3H allele resulted in significantly higher plasma lipid levels than inheritance of the B6 allele. Soat1 variants were also significantly linked to major variations in plasma esterified cholesterol levels but not with free cholesterol levels. Trangenic expression of C3H Soat1 in B6.apoE(-/-) mice resulted in elevations of plasma cholesterol and triglyceride levels. These results indicate that Soat1 is a QTL gene contributing to hyperlipidemia.