Periodic ethanol supply as a path toward unlimited lifespan of Caenorhabditis elegans dauer larvae

The dauer larva is a specialized stage of worm development optimized for survival under harsh conditions that have been used as a model for stress resistance, metabolic adaptations, and longevity. Recent findings suggest that the dauer larva of Caenorhabditis elegans may utilize external ethanol as an energy source to extend their lifespan. It was shown that while ethanol may serve as an effectively infinite source of energy, some toxic compounds accumulating as byproducts of its metabolism may lead to the damage of mitochondria and thus limit the lifespan of larvae. A minimal mathematical model was proposed to explain the connection between the lifespan of a dauer larva and its ethanol metabolism. To explore theoretically if it is possible to extend even further the lifespan of dauer larvae, we incorporated two natural mechanisms describing the recovery of damaged mitochondria and elimination of toxic compounds, which were previously omitted in the model. Numerical simulations of the revised model suggested that while the ethanol concentration is constant, the lifespan still stays limited. However, if ethanol is supplied periodically, with a suitable frequency and amplitude, the dauer could survive as long as we observe the system. Analytical methods further help to explain how feeding frequency and amplitude affect lifespan extension. Based on the comparison of the model with experimental data for fixed ethanol concentration, we proposed the range of feeding protocols that could lead to even longer dauer survival and it can be tested experimentally

Mobilization of cholesterol induces the transition from quiescence to growth in Caenorhabditis elegans through steroid hormone and mTOR signaling

Abstract Recovery from the quiescent developmental stage called dauer is an essential process in C. elegans and provides an excellent model to understand how metabolic transitions contribute to developmental plasticity. Here we show that cholesterol bound to the small secreted proteins SCL-12 or SCL-13 is sequestered in the gut lumen during the dauer state. Upon recovery from dauer, bound cholesterol undergoes endocytosis into lysosomes of intestinal cells, where SCL-12 and SCL-13 are degraded and cholesterol is released. Free cholesterol activates mTORC1 and is used for the production of dafachronic acids. This leads to promotion of protein synthesis and growth, and a metabolic switch at the transcriptional level. Thus, mobilization of sequestered cholesterol stores is the key event for transition from quiescence to growth, and cholesterol is the major signaling molecule in this process

Cytoplasmic condensation induced by membrane damage is associated with antibiotic lethality

AbstractBactericidal antibiotics kill bacteria by perturbing various cellular targets and processes. Disruption of the primary antibiotic-binding partner induces a cascade of molecular events, leading to overproduction of reactive metabolic by-products. It remains unclear, however, how these molecular events contribute to bacterial cell death. Here, we take a single-cell physical biology approach to probe antibiotic function. We show that aminoglycosides and fluoroquinolones induce cytoplasmic condensation through membrane damage and subsequent outflow of cytoplasmic contents as part of their lethality. A quantitative model of membrane damage and cytoplasmic leakage indicates that a small number of nanometer-scale membrane defects in a single bacterium can give rise to the cellular-scale phenotype of cytoplasmic condensation. Furthermore, cytoplasmic condensation is associated with the accumulation of reactive metabolic by-products and lipid peroxidation, and pretreatment of cells with the antioxidant glutathione attenuates cytoplasmic condensation and cell death. Our work expands our understanding of the downstream molecular events that are associated with antibiotic lethality, revealing cytoplasmic condensation as a phenotypic feature of antibiotic-induced bacterial cell death.</jats:p

Systematic Screening for Novel Lipids by Shotgun Lipidomics

A commonly accepted LIPID MAPS classification recognizes eight major lipid categories and over 550 classes, while new lipid classes are still being discovered by targeted biochemical approaches. Despite their compositional diversity, complex lipids such as glycerolipids, glycerophospholipids, saccharolipids, <i>etc.</i> are constructed from unique structural moieties, <i>e.g.</i>, glycerol, fatty acids, choline, phosphate, and trehalose, that are linked by amide, ether, ester, or glycosidic bonds. This modular organization is also reflected in their MS/MS fragmentation pathways, such that common building blocks in different lipid classes tend to generate common fragments. We take advantage of this stereotyped fragmentation to systematically screen for new lipids sharing distant structural similarity to known lipid classes and have developed a discovery approach based on the computational querying of shotgun mass spectra by LipidXplorer software. We applied this concept for screening lipid extracts of <i>C. elegans</i> larvae at the <i>dauer</i> and L3 stages that represent alternative developmental programs executed in response to environmental challenges. The search, covering more than 1.5 million putative chemical compositions, identified a novel class of lyso-maradolipids specifically enriched in <i>dauer</i> larvae

NAD⁺ Is a Food Component That Promotes Exit from Dauer Diapause in <i>Caenorhabditis elegans</i>

<div><p>The free-living soil nematode <i>Caenorhabditis elegans</i> adapts its development to the availability of food. When food is scarce and population density is high, worms enter a developmentally arrested non-feeding diapause stage specialized for long-term survival called the dauer larva. When food becomes available, they exit from the dauer stage, resume growth and reproduction. It has been postulated that compound(s) present in food, referred to as the “food signal”, promote exit from the dauer stage. In this study, we have identified NAD⁺ as a component of bacterial extract that promotes dauer exit. NAD⁺, when dissolved in alkaline medium, causes opening of the mouth and ingestion of food. We also show that to initiate exit from the dauer stage in response to NAD⁺ worms require production of serotonin. Thus, <i>C</i>. <i>elegans</i> can use redox cofactors produced by dietary organisms to sense food.</p></div

Bacterial extract (BE) initiates feeding in <i>C</i>. <i>elegans</i> dauer larva.

<p>(A) Bright field and (B) fluorescent (in shades of yellow) images of dauers incubated in water. (C) Bright field and (D) fluorescent images (in shades of yellow) of dauers incubated in the presence of BE.</p

Endocannabinoids in Caenorhabditis elegans are essential for the mobilization of cholesterol from internal reserves

Proper cholesterol transport is crucial for the functionality of cells. In C. elegans, certain cholesterol derivatives called dafachronic acids (DAs) govern the entry into diapause. In their absence, worms form a developmentally arrested dauer larva. Thus, cholesterol transport to appropriate places for DA biosynthesis warrants the reproductive growth. Recently, we discovered a novel class of glycosphingolipids, PEGCs, required for cholesterol mobilization/transport from internal storage pools. Here, we identify other components involved in this process. We found that strains lacking polyunsaturated fatty acids (PUFAs) undergo increased dauer arrest when grown without cholesterol. This correlates with the depletion of the PUFA-derived endocannabinoids 2-arachidonoyl glycerol and anandamide. Feeding of these endocannabinoids inhibits dauer formation caused by PUFAs deficiency or impaired cholesterol trafficking (e.g. in Niemann-Pick C1 or DAF-7/TGF-β mutants). Moreover, in parallel to PEGCs, endocannabinoids abolish the arrest induced by cholesterol depletion. These findings reveal an unsuspected function of endocannabinoids in cholesterol trafficking regulation

A scheme of dauer larva recovery caused by exposure to the food signal or NAD⁺.

<p>Food signal present in BE is sensed by the dauers, causes opening of the mouth and activation of serotonin signaling, the latter is necessary for pharyngeal pumping. Together, the opening of the mouth and pumping lead to food ingestion and subsequent dauer larva recovery.</p

Active fraction of BE contains NAD⁺.

<p>(A) Mass spectrum of active HPLC fraction (minutes 25–30, ES+). (B) MS/MS of the most abundant ion and elementary composition of the fragments (ES+). (C) Structure of NAD⁺.</p

Food signal requires serotonin to initiate feeding.

<p>(A) Food signal activity of <i>tph-1(n4622)</i> mutant with or without 0.2 mM serotonin. NAD⁺ was at 3 mM concentration. (B) Representative images of the mouth in wild type dauers exposed to borate buffer, (C) BE in water and (E) NAD⁺ in borate buffer. (D) Representative images of the mouth in <i>tph-1(n4622)</i> dauers exposed to BE in water and (F) NAD⁺ in borate buffer. Scale bars show 5 μm in all micrographs. Statistically significant differences are marked with asterisks: p ≤ 0.0001, double asterisk; p ≤ 0.00001, triple asterisk.</p