InteractiveFly: GeneBrief

estrogen-related receptor: Biological Overview | References


Gene name - estrogen-related receptor

Synonyms - dERR

Cytological map position - 66B11-66B11

Function - nuclear receptor

Keywords - A coordinate switch in gene expression that drives a metabolic program normally associated with proliferating cells - supports growth during larval development

Symbol - ERR

FlyBase ID: FBgn0035849

Genetic map position - chr3L:8176477-8179766

Classification - NR_LBD: The ligand binding domain of nuclear receptors; zf-C4: Zinc finger

Cellular location - nuclear



NCBI links: Precomputed BLAST | EntrezGene
Recent literature
Li, H., Chawla, G., Hurlburt, A.J., Sterrett, M.C., Zaslaver, O., Cox, J., Karty, J.A., Rosebrock, A.P., Caudy, A.A. and Tennessen, J.M. (2017). Drosophila larvae synthesize the putative oncometabolite L-2-hydroxyglutarate during normal developmental growth. Proc Natl Acad Sci U S A 114: 1353-1358. PubMed ID: 28115720
Summary:
L-2-hydroxyglutarate (L-2HG) has emerged as a putative oncometabolite that is capable of inhibiting enzymes involved in metabolism, chromatin modification, and cell differentiation. However, despite the ability of L-2HG to interfere with a broad range of cellular processes, this molecule is often characterized as a metabolic waste product. This study demonstrates that Drosophila larvae use the metabolic conditions established by aerobic glycolysis to both synthesize and accumulate high concentrations of L-2HG during normal developmental growth. A majority of the larval L-2HG pool is derived from glucose and dependent on the Drosophila estrogen-related receptor (dERR), which promotes L-2HG synthesis by up-regulating expression of the Drosophila homolog of lactate dehydrogenase (dLdh). dLDH is both necessary and sufficient for directly synthesizing L-2HG and the Drosophila homolog of L-2-hydroxyglutarate dehydrogenase (dL2HGDH), which encodes the enzyme that breaks down L-2HG, is required for stage-specific degradation of the L-2HG pool. In addition, dLDH also indirectly promotes L-2HG accumulation via synthesis of lactate, which activates a metabolic feed-forward mechanism that inhibits dL2HGDH activity and stabilizes L-2HG levels. Finally, dLDH and L-2HG influence position effect variegation and DNA methylation, suggesting that this compound serves to coordinate glycolytic flux with epigenetic modifications. Overall, these data demonstrate that growing animal tissues synthesize L-2HG in a controlled manner, reveal a mechanism that coordinates glucose catabolism with L-2HG synthesis, and establish the fly as a unique model system for studying the endogenous functions of L-2HG during cell growth and proliferation.


BIOLOGICAL OVERVIEW

Metabolism must be coordinated with development to provide the appropriate energetic needs for each stage in the life cycle. Little is known, however, about how this temporal control is achieved. This study shows that the Drosophila ortholog of the estrogen-related receptor (ERR) family of nuclear receptors directs a critical metabolic transition during development. Drosophila ERR mutants die as larvae with low ATP levels and elevated levels of circulating sugars. The expression of active dERR protein in mid-embryogenesis triggers a coordinate switch in gene expression that drives a metabolic program normally associated with proliferating cells, supporting the dramatic growth that occurs during larval development. This study shows that dERR plays a central role in carbohydrate metabolism, demonstrates that a proliferative metabolic program is used in normal developmental growth, and provides a molecular context to understand the close association between mammalian ERR family members and cancer (Tennessen, 2011).

Metabolism must be tightly coupled with developmental progression, with distinct metabolic programs supporting the nutritional and energetic requirements of each stage in the life cycle. Relatively few studies, however, have addressed the molecular mechanisms that provide this temporal coordination. One of the best-characterized metabolic transitions occurs in the developing mammalian heart and is represented by a switch in substrate utilization from glucose and lactate metabolism during fetal stages to postnatal dependence on fatty acid oxidation (Lehman, 2002). This switch is accompanied by the coordinate induction of genes involved in mitochondrial β oxidation and oxidative metabolism and mediated by the nuclear receptors PPARα and ERRγ (Lehman, 2002; Alaynick, 2007). Interestingly, this process reverts in pathological forms of cardiac hypertrophy, demonstrating that metabolic transitions can be associated with disease. In addition, the nutritional status of early developmental stages can have a profound effect on later metabolic health. One of the most dramatic manifestations of this interplay between nutrition and development is the impact of childhood obesity on the incidence of type 2 diabetes and obesity in adults. Obesity also impacts the timing of sexual maturation, linking a metabolic state to a key developmental transition. In spite of this important interplay between nutrition, metabolism, and development, little is understood about how these pathways are integrated (Tennessen, 2011).

Nuclear receptors are a specialized family of ligand-regulated transcription factors that play central roles in controlling development, growth, and metabolism. They are defined by a conserved zinc-finger DNA-binding domain and a C-terminal ligand-binding domain (LBD) that can impart multiple regulatory functions. One subfamily of these receptors are the estrogen-related receptors (ERRs), represented by three paralogs in mammals, ERRα, ERRβ, and ERRγ. Although some synthetic estrogen analogs can suppress the constitutive transcriptional activity of these receptors, they have no known naturally occurring ligands. Genetic studies in mice have demonstrated roles for ERR family members in mitochondrial biogenesis, oxidative phosphorylation, and lipid metabolism (for a review, see Tremblay, 2007). Consistent with these functions, ERRα mutant mice are lean and resistant to diet-induced obesity (Luo, 2003). In addition, ERRα and ERRγ are essential for proper cardiac metabolism. ERRα is required for energy production in response to cardiac stress (Huss, 2007), whereas ERRĪ³ directs a metabolic switch that allows the postnatal heart to metabolize fatty acids (Alaynick, 2007). Recent studies, however, suggest that ERRs play a broader role in metabolic homeostasis (Ao, 2008; Charest-Marcotte, 2010; Eichner, 2010). Moreover, all three mammalian ERRs are associated with cancer progression. ERRα is necessary for the normal growth of estrogen receptor-negative breast cancer tumor grafts, and elevated ERRα expression is associated with aggressive forms of breast cancer. In contrast, ERRβ inhibits the progression of prostate cancer, and increased expression of ERRγ is correlated with a favorable clinical prognosis for breast cance. These observations suggest that ERR family members coordinate cell growth and proliferation with metabolism. The molecular basis for this relationship, however, remains unclear (Tennessen, 2011).

This study presents a functional study of the Drosophila ERR ortholog, dERR. The presence of only a single ERR gene in flies eliminates the potential genetic redundancy between multiple members of the ERR subfamily, allowing determination of its key ancestral functions. dERR is shown to be an essential regulator of carbohydrate metabolism during larval stages. dERR mutants die during the second larval instar with abnormally high levels of circulating sugar and diminished concentrations of ATP and triacylglycerides (TAG). These metabolic defects result from decreased expression of genes involved in glycolysis, the pentose phosphate pathway, and other aspects of carbohydrate metabolism. These genes are coordinately induced midway through embryogenesis, in apparent direct response to the expression of activated dERR protein. Interestingly, this dERR-regulated metabolic state is ideally suited to promote larval growth by converting dietary carbohydrates into biomass and is strikingly reminiscent of the Warburg effect, the observation that cancer cells exhibit glycolysis with lactate secretion and mitochondrial respiration even in the presence of oxygen. These studies reveal that an ERR family member coordinates metabolism with growth, indicate that a proliferative metabolic program is used in the context of normal development, and suggest that mammalian ERRs are associated with cancer through their ability to promote the Warburg effect (Tennessen, 2011).

Drosophila larval development is characterized by an ∼200-fold increase in body mass. This period of dramatic growth requires the efficient conversion of dietary nutrients into cellular building blocks, such as amino acids, fatty acids, and nucleotides. The metabolic basis that supports juvenile growth, however, has not been defined. The studies described here support the model that Drosophila larval growth depends on a form of aerobic glycolysis that is similar to the Warburg effect, and that this metabolic program is established as a mid-embryonic transcriptional switch in response to activated dERR protein (Tennessen, 2011).

Microarray study of dERR mutant larvae revealed a profound and widespread effect on genes that control carbohydrate metabolism, including highly reduced expression of almost all genes in glycolysis and the pentose phosphate pathway (PPP). This is consistent with the ∼2-fold increase in trehalose seen in dERR mutants, along with highly reduced levels of ATP. Mutation of the rate-limiting step in glycolysis, Phosphofructokinase (Pfk), results in elevated trehalose levels, similar to the dERR mutant, and ectopic expression of Phosphoglucose isomerase (Pgi) and Pfk in dERR mutants is sufficient to rescue this phenotype. These studies demonstrate an essential role for dERR in carbohydrate catabolism during larval stages (Tennessen, 2011).

Metabolic profiling confirms these observations, showing significant accumulation of a number of sugars, including glucose-6-phosphate, the first intermediate in the glycolytic pathway. This study, however, also reveals changes in the levels of a number of other key metabolites, providing a broader understanding of dERR function. Lactate is almost completely absent in dERR mutants, consistent with lactate dehydrogenase being one of the most highly downregulated genes in these animals. Levels of α-ketoglutarate, malate, and other late-stage TCA cycle intermediates are also significantly reduced in dERR mutant larvae, along with depletion of several amino acids, of which the most significant and reproducible is proline (Tennessen, 2011).

Interestingly, when taken together with the widespread effects of dERR on glycolysis and the PPP, these changes in metabolite levels are consistent with a form of aerobic glycolysis that is normally associated with cell proliferation. In the context of cancer, this metabolic signature is referred to as the Warburg effect. The increase in carbohydrate metabolism is not designed to produce ATP, but rather promotes the synthesis of amino acids, lipids, and nucleotides, thereby supporting cellular proliferation. Both highly proliferative cells and Drosophila larvae shunt large quantities of glucose through the PPP, allowing them to generate ribose-5-phosphate for nucleotide synthesis and NADPH for fatty acid synthesis and other biosynthetic reactions. Inadequate acetyl-CoA production from glycolysis in dERR mutants and reduced NADPH generation via the PPP likely results in decreased fatty acid synthesis, accounting for the reduced TAG levels in these animals). In support of this hypothesis, TAG levels are reduced by 32% in animals that carry a loss-of-function mutation in glucose-6-phosphate dehydrogenase, which encodes a rate-limiting step in the PPP. Moreover, mutations in PPP enzymes significantly decrease the rate of carbohydrate-dependent fatty acid synthesis in larvae. Similarly, elevated Ldh activity, which is a hallmark of cancer metabolism, is present in normal Drosophila larvae, which display elevated expression of the Drosophila ortholog of Ldh and high levels of Ldh enzyme activity. This enzyme converts pyruvate into lactate, preventing pyruvate from entering the mitochondria and generating NAD+ to promote maximal glycolytic flux. As a result of this diversion of pyruvate away from energy production, the TCA cycle in proliferating cells becomes dependent on amino acids derived from glutamic acid. Large amounts of glutamine are consumed by these cells to anaplerotically maintain the concentration of TCA intermediates. In an analogous manner, Drosophila larval metabolism appears to rely heavily on proline, which is significantly reduced in dERR mutants. Many insects use proline to generate energy and anaplerotically replenish the TCA cycle in the same way that proliferating yeast and cancer cells rely on glutamine. Taken together, these observations support the model that dERR establishes a metabolic state that is related to cellular proliferation, and that this function is essential for larval viability and growth (Tennessen, 2011).

These studies also provide initial insights into the tissue-specific metabolic programs regulated by dERR during larval stages. When dERR is expressed in only the muscle or epidermis, it promotes transcription of the core glycolytic pathway. In contrast, specific expression of dERR in the fat body upregulates Phosphogluconate dehydrogenase (Pgd), which encodes an essential enzyme in the PPP that is induced by sugar consumption. These results are consistent with the metabolic requirements of these different tissues. The dramatic expansion of the epidermis during larval growth, along with its production of cuticle, requires efficient glucose catabolism, as does the muscle to provide larval movement. In contrast, the fat body is one of the principle sites where sugar is processed by the PPP, and efficient lipid storage requires PPP activity. These observations indicate that dERR promotes appropriate tissue-specific metabolic programs during larval development (Tennessen, 2011).

A role for dERR in directing a metabolic state normally associated with cell proliferation provides a new context to understand how Drosophila larvae undergo their remarkable 200-fold increase in mass during the 4 days of larval development. It has long been known that many glycolytic enzymes are induced at the onset of larval development and that the PPP and Ldh are highly active at this stage. No evidence, however, has tied these pathways together. In addition, although yeast use a similar mechanism to support their proliferation, no studies have addressed the metabolic state that accompanies juvenile development in higher organisms. These results suggest that dERR directs a coordinated metabolic program that supports the unusual growth that occurs during this stage. These studies of the dERR-regulated transcriptional program also reveal that this metabolic state is established by a coordinate switch in gene expression during mid-embryogenesis. Accumulation of active dERR protein in 12-18 hr embryos directly induces Pfk transcription and, probably, other key dERR target genes. This transcriptional switch in turn establishes the metabolic requirements for the next stage in development. The timing of the accumulation of active dERR protein could be regulated at a number of levels including posttranslational modifications, ligand binding, and/or cofactor recruitment. Further studies are needed to define the mechanisms that regulate this response. It is interesting to note that the use of aerobic glycolysis to support developmental growth may not be restricted to Drosophila larval stages. Many animals undergo exponential growth during embryonic and fetal stages of development. It will be interesting to determine whether similar metabolic states are associated with early growth in other organisms (Tennessen, 2011).

Studies of mammalian ERR family members have largely focused on their roles in mitochondrial biogenesis and oxidative phosphorylation (for a review, see Tremblay and Giguère, 2007). The microarray study of RNA isolated from dERR mutant larvae, however, identified very few genes associated with β oxidation, the TCA cycle, or the electron transport chain. Although genes involved in oxidoreductase activity are upregulated in dERR mutants, most encode cytochrome P450s with unknown functions. Similarly, a number of genes encoding mitochondrial ribosomal proteins are expressed at reduced levels in dERR mutants, although these effects are relatively minor (≤1.5-fold). dERR mutant larvae have normal mitochondrial genome number and mitochondrial morphology, demonstrating no detectable effect on mitochondrial biogenesis. These observations are consistent with a primary function for dERR in biomass production during larval stages, rather than energy generation and oxidative phosphorylation. Rather, it is speculated that dERR may play a more central role in mitochondrial function during adult stages, when the fly is highly dependent on oxidative metabolism to support its increased mobility (Tennessen, 2011).

Conversely, several recent studies have expanded understanding of the metabolic functions of mammalian ERR family members to include those controlled by dERR. ERRγ regulates glycolytic gene expression in the heart, ERRα is bound to the extended promoters of genes involved in glycolysis and the TCA cycle, and ERRγ regulates several genes in these pathways (Alaynick, 2007; Charest-Marcotte, 2010; Eichner, 2010). Similarly, ERRα is required for the upregulation of genes encoding glycolytic enzymes when cells are raised under hypoxic conditions (Ao, 2008). These studies of dERR define carbohydrate metabolism as a key ancestral function for this nuclear receptor subclass, aspects of which have been conserved through evolution to mammals (Tennessen, 2011).

This work also provides a new context to understand roles for mammalian ERR family members in cancer progression. Many studies have demonstrated a close association between ERR receptors and cancer. The molecular basis for this association, however, remains unclear, likely because of the functional redundancy and crosstalk between mammalian ERR paralogs. A recent paper has provided an initial step in this direction, showing that the miR-378 microRNA, which promotes the Warburg effect in BT-474 cancer cells, downregulates ERRγ expression in this context (Eichner, 2010). These studies of the single Drosophila ERR family member raise the possibility that mammalian ERRs control the dramatic cellular proliferation associated with cancer through their ability to promote the Warburg effect (Tennessen, 2011).

HIF- and non-HIF-regulated hypoxic responses require the estrogen-related receptor in Drosophila melanogaster

Low-oxygen tolerance is supported by an adaptive response that includes a coordinate shift in metabolism and the activation of a transcriptional program that is driven by the hypoxia-inducible factor (HIF) pathway. The precise contribution of HIF-1a in the adaptive response, however, has not been determined. This study investigated how HIF influences hypoxic adaptation throughout Drosophila melanogaster development. Hypoxic-induced transcriptional changes were found to be comprised of HIF-dependent and HIF-independent pathways that are distinct and separable. Normoxic set-points of carbohydrate metabolites are significantly altered in sima mutants and these animals are unable to mobilize glycogen in hypoxia. Furthermore, it was found that the estrogen-related receptor (dERR), which is a global regulator of aerobic glycolysis in larvae, is required for a competent hypoxic response. dERR binds to dHIFa and participates in the HIF-dependent transcriptional program in hypoxia. In addition, dERR acts in the absence of dHIFa in hypoxia and a significant portion of HIF-independent transcriptional responses can be attributed to dERR actions, including upregulation of glycolytic transcripts. These results indicate that competent hypoxic responses arise from complex interactions between HIF-dependent and -independent mechanisms, and that dERR plays a central role in both of these programs (Li, 2013).

A triple mutant of the Drosophila ERR confers ligand-induced suppression of activity

The steroid hormone (NR3) subfamily of nuclear receptors was until recently believed to be restricted to deuterostomes. However, a novel nuclear receptor belonging to the NR3 subfamily was recently identified in the Drosophila genome, indicating the existence of an ancestor before the evolutionary split of deuterostomes and protostomes. This receptor, termed the Drosophila estrogen-related receptor (dERR), most closely resembles the human and mouse estrogen-related receptors (ERRs) in both the DNA binding domain (DBD) (approximately 85% identical) and the ligand binding domain (LBD) (approximately 35% identical). This study describe the functional analysis and rational design of ligand responsive dERR mutants created by protein engineering of the LBD. On the basis of homology modeling, three amino acid residues in the LBD were identified and mutated to enable ligand-dependent suppression of transcriptional activity. The results show that the Y295A/T333I/Y365L triple mutant is significantly suppressed by the known ERR inverse agonists 4-hydroxytamoxifen (OHT) and diethylstilbestrol (DES), in comparison to the wild-type dERR receptor, which was inefficiently suppressed by these substances. The coactivator mGRIP-1 (mouse glucocorticoid receptor interacting protein 1) was shown to significantly increase the activity of the triple mutant in transfection experiments, and the addition of OHT resulted in an efficient suppression of the activity. Accordingly, the ability to functionally interact with a coactivator is still maintained by the Y295A/T333I/Y365L mutant. These findings demonstrate the potential of using rational design and engineering of the LBD to study the function of a nuclear receptor lacking identified ligands (Ostberg, 2003).


REFERENCES

Search PubMed for articles about Drosophila ERR

Alaynick, W. A. et al. (2007). ERRgamma directs and maintains the transition to oxidative metabolism in the postnatal heart. Cell Metab. 6: 13-24. PubMed ID: 17618853

Ao, A., Wang, H., Kamarajugadda, S. and Lu, J. (2000). Involvement of estrogen-related receptors in transcriptional response to hypoxia and growth of solid tumors Proc. Natl. Acad. Sci. 105: 7821-7826. PubMed ID: 18509053

Charest-Marcotte, A. et al. (2010). The homeobox protein Prox1 is a negative modulator of ERRalpha/PGC-1alpha bioenergetic functions Genes Dev. 24: 537-542. PubMed ID: 20194433

Eichner, L. J. et al. (2010). miR-378∗ mediates metabolic shift in breast cancer cells via the PGC-1β/ERRγ transcriptional pathway. Cell Metab. 12: 352-361. PubMed ID: 20889127

Huss, J. M. et al. (2007). The nuclear receptor ERRalpha is required for the bioenergetic and functional adaptation to cardiac pressure overload. Cell Metab. 6: 25-37. PubMed ID: 17618854

Lehman, J. J. and Kelly, D. P. (2002). Transcriptional activation of energy metabolic switches in the developing and hypertrophied heart. Clin. Exp. Pharmacol. Physiol. 29: 339-345. PubMed ID: 11985547

Li, Y., Padmanabha, D., Gentile, L. B., Dumur, C. I., Beckstead, R. B. and Baker, K. D. (2013). HIF- and non-HIF-regulated hypoxic responses require the estrogen-related receptor in Drosophila melanogaster. PLoS Genet 9: e1003230. PubMed ID: 23382692

Luo, J. et al. (2003). Reduced fat mass in mice lacking orphan nuclear receptor estrogen-related receptor alpha. Mol. Cell. Biol. 23: 7947-7956. PubMed ID: 14585956

Ostberg, T. et al. (2003). A triple mutant of the Drosophila ERR confers ligand-induced suppression of activity. Biochemistry 42(21): 6427-35. PubMed ID: 12767224

Tennessen, J. M., Baker, K. D., Lam, G., Evans, J. and Thummel, C. S. (2011). The Drosophila estrogen-related receptor directs a metabolic switch that supports developmental growth. Cell Metab. 13(2): 139-48. PubMed ID: 21284981

Tremblay, A. M. and Giguère, V. (2007). The NR3B subgroup: an overview. Nucl. Recept. Signal. 5: e009. PubMed ID: 18174917


Biological Overview

date revised: 30 April 2015

Home page: The Interactive Fly © 2011 Thomas Brody, Ph.D.

The Interactive Fly resides on the
Society for Developmental Biology's Web server.