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Purpose of Review: Sphingolipids and their metabolites regulate a great variety of cellular processes. Recent findings implicate sphingolipids in the regulation of lipid synthesis, lipoprotein metabolism and the development of atherosclerosis., Recent Findings: Sphingolipid synthesis correlates with the regulation of the sterol-regulatory element-binding proteins - key transcription factors of genes of lipid metabolism. Inhibition of sphingolipid synthesis decreases synthesis of genes regulated by sterol regulatory element-binding protein, such as the rate-limiting enzymes of fatty acid and cholesterol synthesis as well as fatty-acyl-CoA synthases, important in the synthesis of phospholipids. In animal models, inhibition of sphingolipid synthesis correlates with decreased atherosclerotic lesions and a decreased susceptibility of lipoproteins to aggregate--a key mechanism in the development of the atherosclerotic lesion. The demonstration that ceramide and glucosylceramide (metabolites of sphingolipid synthesis) affect cholesterol efflux and mechanisms that regulate plasma high-density lipoprotein concentrations is further evidence for a role of sphingolipids in the regulation of lipid homeostasis. Direct mechanisms of how sphingolipid synthesis regulates lipid synthesis are currently unknown. The recent identification of key proteins of synthesis and specific transport proteins that regulate sphingolipid synthesis, however, is expected to contribute to the understanding about the interdependent regulation of sphingolipid and lipid metabolism., Summary: Emerging data strongly suggest a role of sphingolipid synthesis in the regulation of transcription factors and regulatory proteins that control cellular lipid homeostasis.

Objective: Sterol-regulatory element-binding proteins (SREBPs) regulate transcription of genes of lipid metabolism. Ceramide decreases transcriptionally active SREBP levels independently of intracellular cholesterol levels. Mechanisms of the ceramide-mediated decrease of SREBP levels were investigated., Methods and Results: Experiments were performed in Chinese hamster ovary cells. Inhibition of ceramide synthesis with myriocin, cycloserine, or fumonisin decreases levels of transcriptionally active SREBP and reduces SRE-mediated gene transcription. When ceramide synthesis is increased through exogenous sphingosine or inhibition of sphingosine kinase, SRE-mediated gene transcription is increased. The important role of ceramide synthesis in SRE-mediated gene transcription is confirmed in LY-B cells that do not synthesize ceramide de novo. LY-B cells fail to increase SRE-mediated gene transcription in sterol depletion., Conclusions: Ceramide synthesis correlates with the generation of transcriptionally active SREBP and SRE-mediated gene transcription. Inhibition of ceramide synthesis decreases levels of transcriptionally active SREBP and SRE-mediated gene transcription. It is hypothesized that the process of ongoing ceramide synthesis contributes to the physiological processing of SREBP, perhaps affecting ER-to-Golgi trafficking. Taken together, modification of ceramide synthesis could be a novel target for drug development in the pharmacologic modification of SRE-dependent pathways.

The sterol regulatory element binding-proteins (SREBPs) are transcription factors that regulate the genes of lipid metabolism. Cholesterol and unsaturated fatty acids regulate SREBPs. Giardia lamblia (GL) is an intestinal parasite and one of the earliest derived members within the eukaryotic lineage. GLs exist as trophozoites and cysts. Growth in cholesterol depletion induces transcription of cyst-wall protein (CWP) genes that are upregulated during encystation. The hypothesis was investigated that SREBP-like pathways have a role in cwp gene transcription. Chinese hamster ovary cells were transfected with a cwp-2 promoter reporter construct. Incubation with cholesterol or oleate reduced cwp-2 mediated gene transcription to about half of the control. Incubation in sterol-depleted media, or in the presence of either an inhibitor of intracellular cholesterol movement or inhibitor of cholesterol synthesis, increased gene expression up to 3-fold. Overexpression of SREBPs increased reporter gene activity 2.5-fold. In the absence of functional SREBPs, cwp-2 was not regulated by cholesterol. Footprint analysis of cwp-2 reveals three novel binding sites for mammalian SREBPs with no homologies in other species or humans. The data show that SREBP binds to and can modulate transcription of a regulatory element from an ancient eukaryote and suggest the existence of an SREBP homolog in GL.

A major physiological feedback mechanism of cholesterol in transcription of a number of lipid metabolism-related genes is mediated by sterol regulatory elements (SREs) and their binding proteins (SREBPs). Polyunsaturated free fatty acids alone, as well as synergistically with sterols, decrease SRE-mediated gene expression up to 80% in a dose-dependent manner by decreasing levels of the active transcription factor SREBP. We investigated potential mechanisms for this effect. We hypothesized that free fatty acids reduce SREBP-mediated gene transcription by increasing intracellular cholesterol content through the hydrolysis of cellular sphingomyelin, which has a high affinity for free cholesterol. We also questioned whether the lipid second messenger ceramide, a product of sphingomyelin hydrolysis, can decrease SRE-mediated gene transcription. First we investigated the effect of fatty acids on sphingomyelin hydrolysis. Incubation of [(3)H]choline-labeled cells with unsaturated (but not saturated) fatty acids induced hydrolysis of [(3)H]choline-labeled sphingomyelin. Also, incubation of cell extracts from fatty acid-treated cells with [(3)H]sphingomyelin increased generation of [(3)H]ceramide compared with control cells in vitro. We found that addition of ceramide analogs alone and additively with fatty acids decreased SRE expression and that ceramide analogs reduced levels of the transcriptionally active forms of SREBP-1 and SREBP-2. Increasing intracellular ceramide levels by exogenous sphingomyelinase or inhibition of ceramidase decreased SRE-mediated gene expression. None of the above conditions induced apoptosis. Incubation with U18666A, a compound that inhibits intracellular cholesterol movement, increased SRE-mediated gene transcription. C(2)-ceramide abrogated the effect of U18666A on SRE-mediated gene transcription, suggesting cholesterol-independent regulation of SREBP. We provide evidence that sphingomyelin hydrolysis and intermediates of sphingomyelin metabolism (in addition to cholesterol and fatty acids) contribute to regulation of SRE-mediated gene transcription.