A conserved SREBP-1/phosphatidylcholine feedback circuit regulates lipogenesis in metazoans.

Sterol regulatory element-binding proteins (SREBPs) activate genes involved in the synthesis and trafficking of cholesterol and other lipids and are critical for maintaining lipid homeostasis. Aberrant SREBP activity, however, can contribute to obesity, fatty liver disease, and insulin resistance, hallmarks of metabolic syndrome. Our studies identify a conserved regulatory circuit in which SREBP-1 controls genes in the one-carbon cycle, which produces the methyl donor S-adenosylmethionine (SAMe). Methylation is critical for the synthesis of phosphatidylcholine (PC), a major membrane component, and we find that blocking SAMe or PC synthesis in C. elegans, mouse liver, and human cells causes elevated SREBP-1-dependent transcription and lipid droplet accumulation. Distinct from negative regulation of SREBP-2 by cholesterol, our data suggest a feedback mechanism whereby maturation of nuclear, transcriptionally active SREBP-1 is controlled by levels of PC. Thus, nutritional or genetic conditions limiting SAMe or PC production may activate SREBP-1, contributing to human metabolic disorders.

MicroRNAs in metabolism and metabolic disorders.

MicroRNAs (miRNAs) have recently emerged as key regulators of metabolism. For example, miR-33a and miR-33b have a crucial role in controlling cholesterol and lipid metabolism in concert with their host genes, the sterol-regulatory element-binding protein (SREBP) transcription factors. Other metabolic miRNAs, such as miR-103 and miR-107, regulate insulin and glucose homeostasis, whereas miRNAs such as miR-34a are emerging as key regulators of hepatic lipid homeostasis. The discovery of circulating miRNAs has highlighted their potential as both endocrine signalling molecules and disease markers. Dysregulation of miRNAs may contribute to metabolic abnormalities, suggesting that miRNAs may potentially serve as therapeutic targets for ameliorating cardiometabolic disorders.

A novel 3-hydroxysteroid dehydrogenase that regulates reproductive development and longevity.

Endogenous small molecule metabolites that regulate animal longevity are emerging as a novel means to influence health and life span. In C. elegans, bile acid-like steroids called the dafachronic acids (DAs) regulate developmental timing and longevity through the conserved nuclear hormone receptor DAF-12, a homolog of mammalian sterol-regulated receptors LXR and FXR. Using metabolic genetics, mass spectrometry, and biochemical approaches, we identify new activities in DA biosynthesis and characterize an evolutionarily conserved short chain dehydrogenase, DHS-16, as a novel 3-hydroxysteroid dehydrogenase. Through regulation of DA production, DHS-16 controls DAF-12 activity governing longevity in response to signals from the gonad. Our elucidation of C. elegans bile acid biosynthetic pathways reveals the possibility of novel ligands as well as striking biochemical conservation to other animals, which could illuminate new targets for manipulating longevity in metazoans.

Hormonal control of C. elegans dauer formation and life span by a Rieske-like oxygenase.

C. elegans diapause, gonadal outgrowth, and life span are regulated by a lipophilic hormone, which serves as a ligand to the nuclear hormone receptor DAF-12. A key step in hormone production is catalyzed by the CYP450 DAF-9, but the extent of the biosynthetic pathway is unknown. Here, we identify a conserved Rieske-like oxygenase, DAF-36, as a component in hormone metabolism. Mutants display larval developmental and adult aging phenotypes, as well as patterns of epistasis similar to that of daf-9. Larval phenotypes are potently reversed by crude lipid extracts, 7-dehydrocholesterol, and a recently identified DAF-12 sterol ligand, suggesting that DAF-36 works early in the hormone biosynthetic pathway. DAF-36 is expressed primarily within the intestine, a major organ of metabolic and endocrine control, distinct from DAF-9. These results imply that C. elegans hormone production has multiple steps and is distributed, and that it may provide one way that tissues register their current physiological state during organismal commitments.

MTCH2 is a conserved regulator of lipid homeostasis.

OBJECTIVE: More than one-third of U.S. adults have obesity, causing an alarming increase in obesity-related comorbidities such as type 2 diabetes. The functional role of mitochondrial carrier homolog 2 (MTCH2), a human obesity-associated gene, in lipid homeostasis was investigated in Caenorhabditis elegans, cell culture, and mice. METHODS: In C. elegans, MTCH2/MTCH-1 was depleted, using RNAi and a genetic mutant, and overexpressed to assess its effect on lipid accumulation. In cells and mice, shRNAs against MTCH2 were used for knockdown and MTCH2 overexpression vectors were used for overexpression to study the role of this gene in fat accumulation. RESULTS: MTCH2 knockdown reduced lipid accumulation in adipocyte-like cells in vitro and in C. elegans and mice in vivo. MTCH2 overexpression increased fat accumulation in cell culture, C. elegans, and mice. Acute MTCH2 inhibition reduced fat accumulation in animals subjected to a high-fat diet. Finally, MTCH2 influenced estrogen receptor 1 (ESR1) activity. CONCLUSIONS: MTCH2 is a conserved regulator of lipid homeostasis. MTCH2 was found to be both required and sufficient for lipid homeostasis shifts, suggesting that pharmacological inhibition of MTCH2 could be therapeutic for treatment of obesity and related disorders. MTCH2 could influence lipid homeostasis through inhibition of ESR1 activity.

Control of Caenorhabditis elegans life history by nuclear receptor signal transduction.

Caenorhabditis elegans diapause, reproductive development, and life span are influenced by the DAF-12 nuclear hormone receptor signaling pathway. Here, we describe how this nuclear receptor integrates environmental and physiologic cues and regulates developmental age, reproduction and aging.

FBXO11 promotes ubiquitination of the Snail family of transcription factors in cancer progression and epidermal development.

The Snail family of transcription factors are core inducers of epithelial-to-mesenchymal transition (EMT). Here we show that the F-box protein FBXO11 recognizes and promotes ubiquitin-mediated degradation of multiple Snail family members including Scratch. The association between FBXO11 and Snai1 in vitro is independent of Snai1 phosphorylation. Overexpression of FBXO11 in mesenchymal cells reduces Snail protein abundance and cellular invasiveness. Conversely, depletion of endogenous FBXO11 in epithelial cancer cells causes Snail protein accumulation, EMT, and tumor invasion, as well as loss of estrogen receptor expression in breast cancer cells. Expression of FBXO11 is downregulated by EMT-inducing signals TGFß and nickel. In human cancer, high FBXO11 levels correlate with expression of epithelial markers and favorable prognosis. The results suggest that FBXO11 sustains the epithelial state and inhibits cancer progression. Inactivation of FBXO11 in mice leads to neonatal lethality, epidermal thickening, and increased Snail protein levels in epidermis, validating that FBXO11 is a physiological ubiquitin ligase of Snail. Moreover, in C. elegans, the FBXO11 mutant phenotype is attributed to the Snail factors as it is suppressed by inactivation/depletion of Snail homologs. Collectively, these findings suggest that the FBXO11-Snail regulatory axis is evolutionarily conserved and critically governs carcinoma progression and mammalian epidermal development.

The NHR-8 nuclear receptor regulates cholesterol and bile acid homeostasis in C. elegans.

Hormone-gated nuclear receptors (NRs) are conserved transcriptional regulators of metabolism, reproduction, and homeostasis. Here we show that C. elegans NHR-8 NR, a homolog of vertebrate liver X and vitamin D receptors, regulates nematode cholesterol balance, fatty acid desaturation, apolipoprotein production, and bile acid metabolism. Loss of nhr-8 results in a deficiency in bile acid-like steroids, called the dafachronic acids, which regulate the related DAF-12/NR, thus controlling entry into the long-lived dauer stage through cholesterol availability. Cholesterol supplementation rescues various nhr-8 phenotypes, including developmental arrest, unsaturated fatty acid deficiency, reduced fertility, and shortened life span. Notably, nhr-8 also interacts with daf-16/FOXO to regulate steady-state cholesterol levels and is synthetically lethal in combination with insulin signaling mutants that promote unregulated growth. Our studies provide important insights into nuclear receptor control of cholesterol balance and metabolism and their impact on development, reproduction, and aging in the context of larger endocrine networks.

Pharmacological inhibition of a microRNA family in nonhuman primates by a seed-targeting 8-mer antimiR.

MicroRNAs (miRNAs) regulate many aspects of human biology. They target mRNAs for translational repression or degradation through base pairing with 3' untranslated regions, primarily via seed sequences (nucleotides 2 to 8 in the mature miRNA sequence). A number of individual miRNAs and miRNA families share seed sequences and targets, but differ in the sequences outside of the seed. miRNAs have been implicated in the etiology of a wide variety of human diseases and therefore represent promising therapeutic targets. However, potential redundancy of different miRNAs sharing the same seed sequence and the challenge of simultaneously targeting miRNAs that differ significantly in nonseed sequences complicate therapeutic targeting approaches. We recently demonstrated effective inhibition of entire miRNA families using seed-targeting 8-mer locked nucleic acid (LNA)-modified antimiRs in short-term experiments in mammalian cells and in mice. However, the long-term efficacy and safety of this approach in higher organisms, such as humans and nonhuman primates, have not been determined. We show that pharmacological inhibition of the miR-33 family, key regulators of cholesterol/lipid homeostasis, by a subcutaneously delivered 8-mer LNA-modified antimiR in obese and insulin-resistant nonhuman primates results in derepression of miR-33 targets, such as ABCA1, increases circulating high-density lipoprotein cholesterol, and is well tolerated over 108 days of treatment. These findings demonstrate the efficacy and safety of an 8-mer LNA-antimiR against an miRNA family in a nonhuman primate metabolic disease model, suggesting that this could be a feasible approach for therapeutic targeting of miRNA families sharing the same seed sequence in human diseases.

The Rieske oxygenase DAF-36 functions as a cholesterol 7-desaturase in steroidogenic pathways governing longevity.

Bile acids are cholesterol-derived signaling molecules that regulate mammalian metabolism through sterol-sensing nuclear receptor transcription factors. In C. elegans, bile acid-like steroids called dafachronic acids (DAs) control developmental timing and longevity by activating the nuclear receptor DAF-12. However, little is known about the biosynthesis of these molecules. Here, we show that the DAF-36/Rieske oxygenase works at the first committed step, converting cholesterol to 7-dehydrocholesterol. Its elucidation as a cholesterol 7-desaturase provides crucial biochemical evidence that such oxygenases are key steroidogenic enzymes. By controlling DA production, DAF-36 regulates DAF-12 activities for reproductive development and longevity and may illuminate related pathways in metazoans.

A bile acid-like steroid modulates Caenorhabditis elegans lifespan through nuclear receptor signaling.

Broad aspects of Caenorhabditis elegans life history, including larval developmental timing, arrest at the dauer diapause, and longevity, are regulated by the nuclear receptor DAF-12. Endogenous DAF-12 ligands are 3-keto bile acid-like steroids, called dafachronic acids, which rescue larval defects of hormone-deficient mutants, such as daf-9/cytochrome P450 and daf-36/Rieske oxygenase, and activate DAF-12. Here we examined the effect of dafachronic acid on pathways controlling lifespan. Dafachronic acid supplementation shortened the lifespan of long-lived daf-9 mutants and abolished their stress resistance, indicating that the ligand is "proaging" in response to signals from the dauer pathways. However, the ligand extended the lifespan of germ-line ablated daf-9 and daf-36 mutants, showing that it is "antiaging" in the germ-line longevity pathway. Thus, dafachronic acid regulates C. elegans lifespan according to signaling state. These studies provide key evidence that bile acid-like steroids modulate aging in animals.

Identification of ligands for DAF-12 that govern dauer formation and reproduction in C. elegans.

In response to environmental and dietary cues, the C. elegans orphan nuclear receptor, DAF-12, regulates dauer diapause, reproductive development, fat metabolism, and life span. Despite strong evidence for hormonal control, the identification of the DAF-12 ligand has remained elusive. In this work, we identified two distinct 3-keto-cholestenoic acid metabolites of DAF-9, a cytochrome P450 involved in hormone production, that function as ligands for DAF-12. At nanomolar concentrations, these steroidal ligands (called dafachronic acids) bind and transactivate DAF-12 and rescue the hormone deficiency of daf-9 mutants. Interestingly, DAF-9 has a biochemical activity similar to mammalian CYP27A1 catalyzing addition of a terminal acid to the side chain of sterol metabolites. Together, these results define the first steroid hormones in nematodes as ligands for an invertebrate orphan nuclear receptor and demonstrate that steroidal regulation of reproduction, from biology to molecular mechanism, is conserved from worms to humans.