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1.
Elife ; 122024 Feb 13.
Artículo en Inglés | MEDLINE | ID: mdl-38349720

RESUMEN

Dedicated genetic pathways regulate cysteine homeostasis. For example, high levels of cysteine activate cysteine dioxygenase, a key enzyme in cysteine catabolism in most animal and many fungal species. The mechanism by which cysteine dioxygenase is regulated is largely unknown. In an unbiased genetic screen for mutations that activate cysteine dioxygenase (cdo-1) in the nematode Caenorhabditis elegans, we isolated loss-of-function mutations in rhy-1 and egl-9, which encode proteins that negatively regulate the stability or activity of the oxygen-sensing hypoxia inducible transcription factor (hif-1). EGL-9 and HIF-1 are core members of the conserved eukaryotic hypoxia response. However, we demonstrate that the mechanism of HIF-1-mediated induction of cdo-1 is largely independent of EGL-9 prolyl hydroxylase activity and the von Hippel-Lindau E3 ubiquitin ligase, the classical hypoxia signaling pathway components. We demonstrate that C. elegans cdo-1 is transcriptionally activated by high levels of cysteine and hif-1. hif-1-dependent activation of cdo-1 occurs downstream of an H2S-sensing pathway that includes rhy-1, cysl-1, and egl-9. cdo-1 transcription is primarily activated in the hypodermis where it is also sufficient to drive sulfur amino acid metabolism. Thus, the regulation of cdo-1 by hif-1 reveals a negative feedback loop that maintains cysteine homeostasis. High levels of cysteine stimulate the production of an H2S signal. H2S then acts through the rhy-1/cysl-1/egl-9 signaling pathway to increase HIF-1-mediated transcription of cdo-1, promoting degradation of cysteine via CDO-1.


Proteins are large molecules in our cells that perform various roles, from acting as channels through which nutrients can enter the cell, to forming structural assemblies that help the cell keep its shape. Proteins are formed of chains of building blocks called amino acids. There are 20 common amino acids, each with a different 'side chain' that confers it with specific features. Cysteine is one of these 20 amino acids. Its side chain has a 'thiol' group, made up of a sulfur atom and a hydrogen atom. This thiol group is very reactive, and it is an essential building block of enzymes (proteins that speed up chemical reactions within the cell), structural proteins and signaling molecules. While cysteine is an essential amino acid for the cell to function, excess cysteine can be toxic. The concentration of cysteine in animal cells is tightly regulated by an enzyme called cysteine dioxygenase. This enzyme is implicated in two rare conditions that affect metabolism, where the product of cysteine dioxygenase is a key driver of disease severity. Additionally, cysteine dioxygenase acts as a tumor suppressor gene, and its activity becomes blocked in diverse cancers. Understanding how cysteine dioxygenase is regulated may be important for research into these conditions. While it has been shown that excess cysteine drives the production and activity of cysteine dioxygenase, how the cell detects high levels of cysteine remained unknown. Warnhoff et al. sought to resolve this question using the roundworm Caenorhabditis elegans. First, the scientists demonstrated that, like in mammals, high levels of cysteine drive the production of cysteine dioxygenase in C. elegans. Next, the researchers used an approach called an unbiased genetic screening to find genes that induce cysteine dioxygenase production when they are mutated. These experiments revealed that the protein HIF-1 can drive the production of cysteine dioxygenase when it is activated by a pathway that senses hydrogen sulfide gas. Based on these results, Warnhoff et al. propose that high levels of cysteine lead to the production of hydrogen sulfide gas that in turn drives the production of cysteine dioxygenase via HIF-1 activation of gene expression. The results reported by Warnhoff et al. suggest that modulating HIF-1 signaling could control the activity of cysteine dioxygenase. This information could be used in the future to develop therapies for molybdenum cofactor deficiency, isolated sulfite oxidase deficiency and several types of cancer. However, first it will be necessary to demonstrate that the same signaling pathway is active in humans.


Asunto(s)
Caenorhabditis elegans , Cisteína , Animales , Caenorhabditis elegans/genética , Cisteína-Dioxigenasa/genética , Hipoxia , Factor 1 Inducible por Hipoxia , Homeostasis
2.
bioRxiv ; 2023 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-37205365

RESUMEN

Dedicated genetic pathways regulate cysteine homeostasis. For example, high levels of cysteine activate cysteine dioxygenase, a key enzyme in cysteine catabolism in most animal and many fungal species. The mechanism by which cysteine dioxygenase is regulated is largely unknown. In an unbiased genetic screen for mutations that activate cysteine dioxygenase (cdo-1) in the nematode C. elegans, we isolated loss-of-function mutations in rhy-1 and egl-9, which encode proteins that negatively regulate the stability or activity of the oxygen-sensing hypoxia inducible transcription factor (hif-1). EGL-9 and HIF-1 are core members of the conserved eukaryotic hypoxia response. However, we demonstrate that the mechanism of HIF-1-mediated induction of cdo-1 is largely independent of EGL-9 prolyl hydroxylase activity and the von Hippel-Lindau E3 ubiquitin ligase, the classical hypoxia signaling pathway components. We demonstrate that C. elegans cdo-1 is transcriptionally activated by high levels of cysteine and hif-1. hif-1-dependent activation of cdo-1 occurs downstream of an H2S-sensing pathway that includes rhy-1, cysl-1, and egl-9. cdo-1 transcription is primarily activated in the hypodermis where it is also sufficient to drive sulfur amino acid metabolism. Thus, the regulation of cdo-1 by hif-1 reveals a negative feedback loop that maintains cysteine homeostasis. High levels of cysteine stimulate the production of an H2S signal. H2S then acts through the rhy-1/cysl-1/egl-9 signaling pathway to increase HIF-1-mediated transcription of cdo-1, promoting degradation of cysteine via CDO-1.

3.
J Biol Chem ; 299(1): 102736, 2023 01.
Artículo en Inglés | MEDLINE | ID: mdl-36423681

RESUMEN

Molybdenum cofactor (Moco) is a prosthetic group necessary for the activity of four unique enzymes, including the essential sulfite oxidase (SUOX-1). Moco is required for life; humans with inactivating mutations in the genes encoding Moco-biosynthetic enzymes display Moco deficiency, a rare and lethal inborn error of metabolism. Despite its importance to human health, little is known about how Moco moves among and between cells, tissues, and organisms. The prevailing view is that cells that require Moco must synthesize Moco de novo. Although, the nematode Caenorhabditis elegans appears to be an exception to this rule and has emerged as a valuable system for understanding fundamental Moco biology. C. elegans has the seemingly unique capacity to both synthesize its own Moco as well as acquire Moco from its microbial diet. However, the relative contribution of Moco from the diet or endogenous synthesis has not been rigorously evaluated or quantified biochemically. We genetically removed dietary or endogenous Moco sources in C. elegans and biochemically determined their impact on animal Moco content and SUOX-1 activity. We demonstrate that dietary Moco deficiency dramatically reduces both animal Moco content and SUOX-1 activity. Furthermore, these biochemical deficiencies have physiological consequences; we show that dietary Moco deficiency alone causes sensitivity to sulfite, the toxic substrate of SUOX-1. Altogether, this work establishes the biochemical consequences of depleting dietary Moco or endogenous Moco synthesis in C. elegans and quantifies the surprising contribution of the diet to maintaining Moco homeostasis in C. elegans.


Asunto(s)
Metaloproteínas , Cofactores de Molibdeno , Sulfito-Oxidasa , Animales , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Dieta , Metaloproteínas/genética , Metaloproteínas/metabolismo , Molibdeno/metabolismo , Cofactores de Molibdeno/metabolismo , Pteridinas/metabolismo , Sulfito-Oxidasa/genética , Sulfito-Oxidasa/metabolismo
4.
MicroPubl Biol ; 20222022.
Artículo en Inglés | MEDLINE | ID: mdl-35224462

RESUMEN

Molybdenum cofactor (Moco) is an essential prosthetic group that mediates the activity of 4 animal oxidases and is required for viability. Humans with mutations in the genes encoding Moco-biosynthetic enzymes suffer from Moco deficiency, a neonatal lethal inborn error of metabolism. Caenorhabditis elegans has recently emerged as a useful and tractable genetic discovery engine for Moco biology. Here, we identify and characterize K10D2.7/moc-6, the C. elegans ortholog of human MOCS2A, a sulfur-carrier protein essential for Moco synthesis. Using CRISPR/Cas9 gene editing, we generate 3 null mutations in K10D2.7/moc-6 and with these alleles genetically demonstrate that K10D2.7/moc-6 is necessary for endogenous Moco synthesis in C. elegans.

5.
Proc Natl Acad Sci U S A ; 118(42)2021 10 19.
Artículo en Inglés | MEDLINE | ID: mdl-34649987

RESUMEN

Cadmium is an environmental pollutant and significant health hazard that is similar to the physiological metal zinc. In Caenorhabditis elegans, high zinc homeostasis is regulated by the high zinc activated nuclear receptor (HIZR-1) transcription factor. To define relationships between the responses to high zinc and cadmium, we analyzed transcription. Many genes were activated by both high zinc and cadmium, and hizr-1 was necessary for activation of a subset of these genes; in addition, many genes activated by cadmium did not require hizr-1, indicating there are at least two mechanisms of cadmium-regulated transcription. Cadmium directly bound HIZR-1, promoted nuclear accumulation of HIZR-1 in intestinal cells, and activated HIZR-1-mediated transcription via the high zinc activation (HZA) enhancer. Thus, cadmium binding promotes HIZR-1 activity, indicating that cadmium acts as a zinc mimetic to hijack the high zinc response. To elucidate the relationships between high zinc and cadmium detoxification, we analyzed genes that function in three pathways: the pcs-1/phytochelatin pathway strongly promoted cadmium resistance but not high zinc resistance, the hizr-1/HZA pathway strongly promoted high zinc resistance but not cadmium resistance, and the mek-1/sek-1/kinase signaling pathway promoted resistance to high zinc and cadmium. These studies identify resistance pathways that are specific for high zinc and cadmium, as well as a shared pathway.


Asunto(s)
Cadmio/farmacología , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/efectos de los fármacos , Receptores Citoplasmáticos y Nucleares/metabolismo , Zinc/metabolismo , Animales , Caenorhabditis elegans/metabolismo , Homeostasis , Ligandos , Metalotioneína/metabolismo , Unión Proteica
6.
Genes Dev ; 35(3-4): 212-217, 2021 02 01.
Artículo en Inglés | MEDLINE | ID: mdl-33446569

RESUMEN

The molybdenum cofactor (Moco) is a 520-Da prosthetic group that is synthesized in all domains of life. In animals, four oxidases (among them sulfite oxidase) use Moco as a prosthetic group. Moco is essential in animals; humans with mutations in genes that encode Moco biosynthetic enzymes display lethal neurological and developmental defects. Moco supplementation seems a logical therapy; however, the instability of Moco has precluded biochemical and cell biological studies of Moco transport and bioavailability. The nematode Caenorhabditis elegans can take up Moco from its bacterial diet and transport it to cells and tissues that express Moco-requiring enzymes, suggesting a system for Moco uptake and distribution. Here we show that protein-bound Moco is the stable, bioavailable species of Moco taken up by C. elegans from its diet and is an effective dietary supplement, rescuing a Celegans model of Moco deficiency. We demonstrate that diverse Moco:protein complexes are stable and bioavailable, suggesting a new strategy for the production and delivery of therapeutically active Moco to treat human Moco deficiency.


Asunto(s)
Caenorhabditis elegans/metabolismo , Coenzimas/administración & dosificación , Errores Innatos del Metabolismo de los Metales/terapia , Metaloproteínas/administración & dosificación , Pteridinas/administración & dosificación , Animales , Bacterias/metabolismo , Transporte Biológico , Coenzimas/deficiencia , Coenzimas/farmacocinética , Humanos , Metaloproteínas/deficiencia , Metaloproteínas/farmacocinética , Cofactores de Molibdeno , Unión Proteica , Pteridinas/farmacocinética
7.
Nat Chem Biol ; 15(5): 480-488, 2019 05.
Artículo en Inglés | MEDLINE | ID: mdl-30911177

RESUMEN

The kingdoms of life share many small molecule cofactors and coenzymes. Molybdenum cofactor (Moco) is synthesized by many archaea, bacteria, and eukaryotes, and is essential for human development. The genome of Caenorhabditis elegans contains all of the Moco biosynthesis genes, and surprisingly these genes are not essential if the animals are fed a bacterial diet that synthesizes Moco. C. elegans lacking both endogenous Moco synthesis and dietary Moco from bacteria arrest development, demonstrating interkingdom Moco transfer. Our screen of Escherichia coli mutants identifies genes necessary for synthesis of bacterial Moco or transfer to C. elegans. Developmental arrest of Moco-deficient C. elegans is caused by loss of sulfite oxidase, a Moco-requiring enzyme, and is suppressed by mutations in either C. elegans cystathionine gamma-lyase or cysteine dioxygenase, blocking toxic sulfite production from cystathionine. Thus, we define the genetic pathways for an interkingdom dialogue focused on sulfur homeostasis.


Asunto(s)
Caenorhabditis elegans/metabolismo , Coenzimas/metabolismo , Metaloproteínas/metabolismo , Pteridinas/metabolismo , Sulfitos/metabolismo , Animales , Caenorhabditis elegans/efectos de los fármacos , Homeostasis , Cofactores de Molibdeno , Sulfitos/toxicidad
8.
PLoS Biol ; 16(6): e2005069, 2018 06.
Artículo en Inglés | MEDLINE | ID: mdl-29879108

RESUMEN

Sperm activation is a fascinating example of cell differentiation, in which immotile spermatids undergo a rapid and dramatic transition to become mature, motile sperm. Because the sperm nucleus is transcriptionally silent, this transition does not involve transcriptional changes. Although Caenorhabditis elegans is a leading model for studies of sperm activation, the mechanisms by which signaling pathways induce this transformation remain poorly characterized. Here we show that a conserved transmembrane zinc transporter, ZIPT-7.1, regulates the induction of sperm activation in Caenorhabditis nematodes. The zipt-7.1 mutant hermaphrodites cannot self-fertilize, and males reproduce poorly, because mutant spermatids are defective in responding to activating signals. The zipt-7.1 gene is expressed in the germ line and functions in germ cells to promote sperm activation. When expressed in mammalian cells, ZIPT-7.1 mediates zinc transport with high specificity and is predominantly located on internal membranes. Finally, genetic epistasis places zipt-7.1 at the end of the spe-8 sperm activation pathway, and ZIPT-7.1 binds SPE-4, a presenilin that regulates sperm activation. Based on these results, we propose a new model for sperm activation. In spermatids, inactive ZIPT-7.1 is localized to the membranous organelles, which contain higher levels of zinc than the cytoplasm. When sperm activation is triggered, ZIPT-7.1 activity increases, releasing zinc from internal stores. The resulting increase in cytoplasmic zinc promotes the phenotypic changes characteristic of activation. Thus, zinc signaling is a key step in the signal transduction process that mediates sperm activation, and we have identified a zinc transporter that is central to this activation process.


Asunto(s)
Proteínas de Caenorhabditis elegans/fisiología , Caenorhabditis elegans/fisiología , Proteínas Portadoras/fisiología , Espermatogénesis/fisiología , Secuencia de Aminoácidos , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas Portadoras/genética , Epistasis Genética , Femenino , Genes de Helminto , Transporte Iónico , Masculino , Proteínas de la Membrana/metabolismo , Modelos Biológicos , Mutación , Filogenia , Transducción de Señal , Espermátides/metabolismo , Espermatocitos/metabolismo , Espermatogénesis/genética , Espermatozoides/metabolismo , Zinc/metabolismo
9.
PLoS Biol ; 15(1): e2000094, 2017 01.
Artículo en Inglés | MEDLINE | ID: mdl-28095401

RESUMEN

Nuclear receptors were originally defined as endocrine sensors in humans, leading to the identification of the nuclear receptor superfamily. Despite intensive efforts, most nuclear receptors have no known ligand, suggesting new ligand classes remain to be discovered. Furthermore, nuclear receptors are encoded in the genomes of primitive organisms that lack endocrine signaling, suggesting the primordial function may have been environmental sensing. Here we describe a novel Caenorhabditis elegans nuclear receptor, HIZR-1, that is a high zinc sensor in an animal and the master regulator of high zinc homeostasis. The essential micronutrient zinc acts as a HIZR-1 ligand, and activated HIZR-1 increases transcription of genes that promote zinc efflux and storage. The results identify zinc as the first inorganic molecule to function as a physiological ligand for a nuclear receptor and direct environmental sensing as a novel function of nuclear receptors.


Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Homeostasis/efectos de los fármacos , Receptores Citoplasmáticos y Nucleares/metabolismo , Zinc/farmacología , Animales , Caenorhabditis elegans/efectos de los fármacos , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/química , Proteínas de Caenorhabditis elegans/genética , Proteínas de Transporte de Catión/genética , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , ADN/metabolismo , Elementos de Facilitación Genéticos/genética , Retroalimentación Fisiológica/efectos de los fármacos , Pruebas Genéticas , Mucosa Intestinal/metabolismo , Intestinos/efectos de los fármacos , Ligandos , Mutación/genética , Regiones Promotoras Genéticas/genética , Dominios Proteicos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores Citoplasmáticos y Nucleares/química , Receptores Citoplasmáticos y Nucleares/genética , Transcripción Genética/efectos de los fármacos
10.
Worm ; 4(2): e1023498, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26435887

RESUMEN

Protein N-terminal acetylation is a widespread posttranslational modification in eukaryotes that is catalyzed by N-terminal acetyltransferases (NATs). The biochemical activity of NATs has been characterized extensively, whereas the biological function of NATs is only beginning to be defined. Here we comment on recent progress in understanding the function of NAT activity in C. elegans based on the characterization of natc-1 by Warnhoff et al. (2014) and daf-31 by Chen et al. (2014).(1,2) natc-1 encodes an auxiliary subunit of the NatC complex and modulates stress tolerance, dauer entry, and adult lifespan. daf-31 encodes the catalytic subunit of the NatA complex and affects dauer entry, dauer formation, and adult lifespan. The analysis of these genes and genetic studies of NATs in other organisms suggests protein N-terminal acetylation plays an evolutionarily conserved role in promoting growth and development and inhibiting stress resistance. Furthermore, we propose that NATs may regulate growth and development in response to external cues such as nutrient deprivation and other physiologic stresses.

11.
Nucleic Acids Res ; 43(2): 803-16, 2015 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25552416

RESUMEN

Zinc is essential for biological systems, and aberrant zinc metabolism is implicated in a broad range of human diseases. To maintain homeostasis in response to fluctuating levels of dietary zinc, animals regulate gene expression; however, mechanisms that mediate the transcriptional response to fluctuating levels of zinc have not been fully defined. Here, we identified DNA enhancer elements that mediate intestine-specific transcriptional activation in response to high levels of dietary zinc in C. elegans. Using bioinformatics, we characterized an evolutionarily conserved enhancer element present in multiple zinc-inducible genes, the high zinc activation (HZA) element. The HZA was consistently adjacent to a GATA element that mediates expression in intestinal cells. Functional studies using transgenic animals demonstrated that this modular system of DNA enhancers mediates tissue-specific transcriptional activation in response to high levels of dietary zinc. We used this information to search the genome and successfully identified novel zinc-inducible genes. To characterize the mechanism of enhancer function, we demonstrated that the GATA transcription factor ELT-2 and the mediator subunit MDT-15 are necessary for zinc-responsive transcriptional activation. These findings define new mechanisms of zinc homeostasis and tissue-specific regulation of transcription.


Asunto(s)
Elementos de Facilitación Genéticos , Activación Transcripcional , Zinc/farmacología , Animales , Secuencia de Bases , Cadmio/farmacología , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Secuencia Conservada , ADN de Helmintos/química , Factores de Transcripción GATA/metabolismo , Especificidad de Órganos , Regiones Promotoras Genéticas , Factores de Transcripción/metabolismo , Activación Transcripcional/efectos de los fármacos
12.
PLoS Genet ; 10(10): e1004703, 2014 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-25330323

RESUMEN

The insulin/IGF-1 signaling pathway plays a critical role in stress resistance and longevity, but the mechanisms are not fully characterized. To identify genes that mediate stress resistance, we screened for C. elegans mutants that can tolerate high levels of dietary zinc. We identified natc-1, which encodes an evolutionarily conserved subunit of the N-terminal acetyltransferase C (NAT) complex. N-terminal acetylation is a widespread modification of eukaryotic proteins; however, relatively little is known about the biological functions of NATs. We demonstrated that loss-of-function mutations in natc-1 cause resistance to a broad-spectrum of physiologic stressors, including multiple metals, heat, and oxidation. The C. elegans FOXO transcription factor DAF-16 is a critical target of the insulin/IGF-1 signaling pathway that mediates stress resistance, and DAF-16 is predicted to directly bind the natc-1 promoter. To characterize the regulation of natc-1 by DAF-16 and the function of natc-1 in insulin/IGF-1 signaling, we analyzed molecular and genetic interactions with key components of the insulin/IGF-1 pathway. natc-1 mRNA levels were repressed by DAF-16 activity, indicating natc-1 is a physiological target of DAF-16. Genetic studies suggested that natc-1 functions downstream of daf-16 to mediate stress resistance and dauer formation. Based on these findings, we hypothesize that natc-1 is directly regulated by the DAF-16 transcription factor, and natc-1 is a physiologically significant effector of the insulin/IGF-1 signaling pathway that mediates stress resistance and dauer formation. These studies identify a novel biological function for natc-1 as a modulator of stress resistance and dauer formation and define a functionally significant downstream effector of the insulin/IGF-1 signaling pathway. Protein N-terminal acetylation mediated by the NatC complex may play an evolutionarily conserved role in regulating stress resistance.


Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiología , Factores de Transcripción Forkhead/metabolismo , Insulina/metabolismo , Acetiltransferasa C N-Terminal/metabolismo , Estrés Fisiológico , Acetilación , Acetiltransferasas , Secuencia de Aminoácidos , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/efectos de los fármacos , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Factores de Transcripción Forkhead/genética , Regulación del Desarrollo de la Expresión Génica , Factor I del Crecimiento Similar a la Insulina/metabolismo , Datos de Secuencia Molecular , Mutación , Acetiltransferasa C N-Terminal/genética , Transducción de Señal , Zinc/metabolismo , Zinc/toxicidad
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