Zinc is essential for many cellular processes. To use Caenorhabditis elegans to study zinc metabolism, we developed culture conditions allowing full control of dietary zinc and methods to measure zinc content of animals. Dietary zinc dramatically affected growth and zinc content; wild-type worms survived from 7 mm to 1.3 mm dietary zinc, and zinc content varied 27-fold. We investigated cdf-2, which encodes a predicted zinc transporter in the cation diffusion facilitator family. cdf-2 mRNA levels were increased by high dietary zinc, suggesting cdf-2 promotes zinc homeostasis. CDF-2 protein was expressed in intestinal cells and localized to cytosolic vesicles. A cdf-2 loss-of-function mutant displayed impaired growth and reduced zinc content, indicating that CDF-2 stores zinc by transport into the lumen of vesicles. The relationships between three cdf genes, cdf-1, cdf-2, and sur-7, were analyzed in double and triple mutant animals. A cdf-1 mutant displayed increased zinc content, whereas a cdf-1 cdf-2 double mutant had intermediate zinc content, suggesting cdf-1 and cdf-2 have antagonistic functions. These studies advance C. elegans as a model of zinc metabolism and identify cdf-2 as a new gene that has a critical role in zinc storage.

Graphical AbstractHighlights d phm-2 encodes a SAFB protein conserved in mammals d phm-2 and eat-2 mutants have a defective pharynx and accumulate bacteria in the gut d Bacterial accumulation activates innate immunity and bacterial avoidance behavior d Bacterial avoidance leads to dietary restriction and delays agingIn Brief Kumar et al. link immunity and aging in C. elegans. The authors characterize phm-2 and eat-2, mutants that allow live bacteria to accumulate in the intestine, causing delayed reproductive and somatic aging. The mechanism combines molecular immune activation and behavioral food avoidance, leading to dietary restriction and extended lifespan. SUMMARYMechanisms that control aging are important yet poorly defined. To discover longevity control genes, we performed a forward genetic screen for delayed reproductive aging in C. elegans. Here, we show that am117 is a nonsense mutation in the phm-2 gene, which encodes a protein homologous to human scaffold attachment factor B. phm-2(lf) mutant worms have an abnormal pharynx grinder, which allows live bacteria to accumulate in the intestine. This defect shortens lifespan on highly pathogenic bacteria but extends lifespan and health span on the standard E. coli diet by activating innate immunity pathways that lead to bacterial avoidance behavior and dietary restriction. eat-2(lf) mutants displayed a similar phenotype, indicating accumulation of live bacteria also triggers extended longevity in this mutant. The analysis of phm-2 elucidates connections between pathogen response and aging by defining a mechanism of longevity extension in C. elegans-bacterial colonization, innate immune activation, and bacterial avoidance behavior.

Zinc plays many critical roles in biological systems: zinc bound to proteins has structural and catalytic functions, and zinc is proposed to act as a signaling molecule. Because zinc deficiency and excess result in toxicity, animals have evolved sophisticated mechanisms for zinc metabolism and homeostasis. However, these mechanisms remain poorly defined. To identify genes involved in zinc metabolism, we conducted a forward genetic screen for chemically induced mutations that cause Caenorhabditis elegans to be resistant to high levels of dietary zinc. Nineteen mutations that confer significant resistance to supplemental dietary zinc were identified. To determine the map positions of these mutations, we developed a genomewide map of single nucleotide polymorphisms (SNPs) that can be scored by the high-throughput method of DNA pyrosequencing. This map was used to determine the approximate chromosomal position of each mutation, and the accuracy of this approach was verified by conducting three-factor mapping experiments with mutations that cause visible phenotypes. This is a generally applicable mapping approach that can be used to position a wide variety of C. elegans mutations. The mapping experiments demonstrate that the 19 mutations identify at least three genes that, when mutated, confer resistance to toxicity caused by supplemental dietary zinc. These genes are likely to be involved in zinc metabolism, and the analysis of these genes will provide insights into mechanisms of excess zinc toxicity.

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.

Ethosuximide is a medication used to treat seizure disorders in humans, and we previously demonstrated that ethosuximide can delay age-related changes and extend the lifespan of the nematode Caenorhabditis elegans. The mechanism of action of ethosuximide in lifespan extension is unknown, and elucidating how ethosuximide functions is important for defining endogenous processes that influence lifespan and for exploring the potential of ethosuximide as a therapeutic for age-related diseases. To identify genes that mediate the activity of ethosuximide, we conducted a genetic screen and identified mutations in two genes, che-3 and osm-3, that cause resistance to ethosuximide-mediated toxicity. Mutations in che-3 and osm-3 cause defects in overlapping sets of chemosensory neurons, resulting in defective chemosensation and an extended lifespan. These findings suggest that ethosuximide extends lifespan by inhibiting the function of specific chemosensory neurons. This model is supported by the observation that ethosuximide-treated animals displayed numerous phenotypic similarities with mutants that have chemosensory defects, indicating that ethosuximide inhibits chemosensory function. Furthermore, ethosuximide extends lifespan by inhibiting chemosensation, since the long-lived osm-3 mutants were resistant to the lifespan extension caused by ethosuximide. These studies demonstrate a novel mechanism of action for a lifespan-extending drug and indicate that sensory perception has a critical role in controlling lifespan. Sensory perception also influences the lifespan of Drosophila, suggesting that sensory perception has an evolutionarily conserved role in lifespan control. These studies highlight the potential of ethosuximide and related drugs that modulate sensory perception to extend lifespan in diverse animals.