OBJECTIVE: To investigate the changes of lipid metabolism and stress response of adult C.elegans exposed to non-freezing low temperature and explore the possible mechanism. METHODS: The survival rate and activity of adult C.elegans cultured at 20℃ or 4℃ were observed.Lipid metabolism of the cultured adult C.elegans was evaluated using oil red O staining and by detecting the expressions of the genes related with lipid metabolism.The effects of low temperature exposure on stress level of adult C.elegans were evaluated using mitochondrial fluorescence staining and by detecting the expression levels of stress-related genes and antioxidant genes at both the mRNA and protein levels. RESULTS: The lifespan and activity of adult C.elegans exposed to low temperature were significantly reduced with decreased lipid accumulation (P < 0.05) and decreased expressions of genes related with fatty acid synthesis and metabolism (fat-5, fat-6, fat-7, fasn-1, nhr-49, acs-2 and aco-1;P < 0.01).Cold stress significantly increased the expressions of heat shock proteins hsp-70 and hsp16.2(P < 0.05) but lowered the number of mitochondria (P < 0.0001) and the expressions of atfs-1, sod-2, sod-3 and gpx-1(P < 0.05).Knockout of fat-5, nhr-49 or both fat-5 and fat-6 obviously enhanced the sensitivity of C.elegans to cold stress as shown by further reduced activity (P < 0.05) and reduced survival rate at 24 h (P < 0.0001) under cold stress. CONCLUSION Exposure to a low temperature at 4℃ results in lowered lipid metabolism of adult C.elegans accompanied by a decreased mitochondrial number and quality control ability, which triggers high expressions of stress-related genes and causes reduction of antioxidant capacity, thus callsing lowered activity and reduced lifespan of C.elegans.
Animals ; Antioxidants/metabolism* ; Caenorhabditis elegans ; Caenorhabditis elegans Proteins/genetics* ; Cold-Shock Response ; Lipid Metabolism ; Lipid Metabolism Disorders ; Longevity/genetics*

Chloride channels belong to a superfamily of ion channels that permit passive passage of anions, mainly chloride, across cell membrane. They play a variety of important physiological roles in regulation of cytosolic pH, cell volume homeostasis, organic solute transport, cell migration, cell proliferation, and differentiation. However, little is known about the functional regulation of these channels. In this study, we generated an integrated transgenic worm strain expressing green fluorescence protein (GFP) fused CLC-type chloride channel 1 (CLH-1::GFP), a voltage-gated chloride channel in Caenorhabditis elegans (C. elegans). CLH-1::GFP was expressed in some unidentified head neurons and posterior intestinal cells of C. elegans. Interacting proteins of CLH-1::GFP were purified by GFP-Trap, a novel system for efficient isolation of GFP fusion proteins and their interacting factors. Mass spectrometry (MS) analysis revealed that a total of 27 high probability interacting proteins were co-trapped with CLHp-1::GFP. Biochemical evidence showed that eukaryotic translation elongation factor 1 (EEF-1), one of these co-trapped proteins identified by MS, physically interacted with CLH-1, in consistent with GFP-Trap experiments. Further immunostaining data revealed that the protein level of CLH-1 was significantly increased upon co-expression with EEF-1. These results suggest that the combination of GFP-Trap purification with MS is an excellent tool to identify novel interacting proteins of voltage-gated chloride channels in C. elegans. Our data also show that EEF-1 is a regulator of voltage-gated chloride channel CLH-1.
Animals ; Animals, Genetically Modified ; Caenorhabditis elegans ; genetics ; metabolism ; Caenorhabditis elegans Proteins ; metabolism ; Chloride Channels ; metabolism ; Green Fluorescent Proteins ; chemistry ; Mass Spectrometry ; Peptide Elongation Factor 1 ; metabolism

Adaptation, Physiological ; genetics ; Animals ; Body Temperature ; physiology ; Caenorhabditis elegans ; genetics ; metabolism ; physiology ; Caenorhabditis elegans Proteins ; metabolism ; Gene Expression Regulation ; Humans ; Transcription Factors ; metabolism

Reproduction, fat metabolism, and longevity are intertwined regulatory axes; recent studies in C. elegans have provided evidence that these processes are directly coupled. However, the mechanisms by which they are coupled and the reproductive signals modulating fat metabolism and lifespan are poorly understood. Here, we find that an oogenesis-enriched gene, c30f12.4, is specifically expressed and located in germ cells and early embryos; when the gene is knocked out, oogenesis is disrupted and brood size is decreased. In addition to the reproductive phenotype, we find that the loss of c30f12.4 alters fat metabolism, resulting in decreased fat storage and smaller lipid droplets. Meanwhile, c30f12.4 mutant worms display a shortened lifespan. Our results highlight an important role for c30f12.4 in regulating reproduction, fat homeostasis, and aging in C. elegans, which helps us to better understand the relationship between these processes.
Animals ; Caenorhabditis elegans ; genetics ; metabolism ; Caenorhabditis elegans Proteins ; genetics ; metabolism ; Female ; Lipid Droplets ; metabolism ; Lipid Metabolism ; physiology ; Longevity ; physiology ; Mutation ; Oogenesis ; physiology

OBJECTIVETo explore the expression changes of age-related genes in different stages of aging and the regulating effects of Chuanxiong extract on it.

METHODAccording to the different stages of aging, the experiments were tested at two time points of 2 d and 6 d. Using realtime RT-PCR (qRT-PCR) to test the expression change of aging-related genes among the groups.

RESULTCompared with the 2 d control group, the expression of age-1, daf-2, let-363 were up-regulated in the 6 d control group (P < 0.05) while the expression of ins-18, let-60, sir-2.1, sod-3 were down-regulated (P < 0.05). Compared with the 2 d administration group, the expression of age-1, daf-2, let-363 were significantly up-regulated (P < 0.01) in the 6 d administration group after treated with CXE while the expression of ins-18, let-60, sir-2.1, sod-3 were significantly down-regulated (P < 0.01).

CONCLUSIONIn the progress of aging, the expression of age-1, daf-2, let-363 increased, functioning as aging-promoting genes; while the expression of ins-18, let-60, sir-2.1, sod-3 decreased, functioning as longevity genes; CXE extended the lifespan through inhibiting the expression of these aging-promoting genes and increasing the expression of longevity genes, which would be the molecular mechaniSm of anti-aging of traditional Chinese medicine that can promote Qi and activate blood.

Animals ; Caenorhabditis elegans ; genetics ; growth & development ; metabolism ; Caenorhabditis elegans Proteins ; genetics ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Gene Expression Regulation, Developmental ; drug effects ; Longevity ; drug effects

In this study, we aimed to evaluate the toxic effects, changes in life span, and expression of various metabolism-related genes in Caenorhabditis elegans, using RNA interference (RNAi) and mutant strains, after 3-bromopyruvate (3-BrPA) treatment. C. elegans was treated with various concentrations of 3-BrPA on nematode growth medium (NGM) plates, and their survival was monitored every 24 h. The expression of genes related to metabolism was measured by the real-time fluorescent quantitative polymerase chain reaction (qPCR). Nematode survival in the presence of 3-BrPA was also studied after silencing three hexokinase (HK) genes. The average life span of C. elegans cultured on NGM with 3-BrPA was shortened to 5.7 d compared with 7.7 d in the control group. hxk-1, hxk-2, and hxk-3 were overexpressed after the treatment with 3-BrPA. After successfully interfering hxk-1, hxk-2, and hxk-3, the 50% lethal concentration (LC₅₀) of all mutant nematodes decreased with 3-BrPA treatment for 24 h compared with that of the control. All the cyp35 genes tested were overexpressed, except cyp-35B3. The induction of cyp-35A1 expression was most obvious. The LC₅₀ values of the mutant strains cyp-35A1, cyp-35A2, cyp-35A4, cyp-35B3, and cyp-35C1 were lower than that of the control. Thus, the toxicity of 3-BrPA is closely related to its effect on hexokinase metabolism in nematodes, and the cyp-35 family plays a key role in the metabolism of 3-BrPA.
Animals ; Caenorhabditis elegans/metabolism* ; Caenorhabditis elegans Proteins/genetics* ; Cytochrome P-450 Enzyme System/genetics* ; Hexokinase/physiology* ; Pyruvates/toxicity* ; RNA, Messenger/analysis*

Zn²⁺ is required for the activity of many mitochondrial proteins, which regulate mitochondrial dynamics, apoptosis and mitophagy. However, it is not understood how the proper mitochondrial Zn²⁺ level is achieved to maintain mitochondrial homeostasis. Using Caenorhabditis elegans, we reveal here that a pair of mitochondrion-localized transporters controls the mitochondrial level of Zn²⁺. We demonstrate that SLC-30A9/ZnT9 is a mitochondrial Zn²⁺ exporter. Loss of SLC-30A9 leads to mitochondrial Zn²⁺ accumulation, which damages mitochondria, impairs animal development and shortens the life span. We further identify SLC-25A25/SCaMC-2 as an important regulator of mitochondrial Zn²⁺ import. Loss of SLC-25A25 suppresses the abnormal mitochondrial Zn²⁺ accumulation and defective mitochondrial structure and functions caused by loss of SLC-30A9. Moreover, we reveal that the endoplasmic reticulum contains the Zn²⁺ pool from which mitochondrial Zn²⁺ is imported. These findings establish the molecular basis for controlling the correct mitochondrial Zn²⁺ levels for normal mitochondrial structure and functions.
Animals ; Caenorhabditis elegans/metabolism* ; Cation Transport Proteins/genetics* ; Homeostasis ; Mitochondria/metabolism* ; Zinc/metabolism*

Lipid droplets, which are conserved across almost all species, are cytoplasmic organelles used to store neutral lipids. Identification of lipid droplet regulators will be conducive to resolving obesity and other fat-associated diseases. In this paper, we selected 11 candidates that might be associated with lipid metabolism in Caenorhabditis elegans. Using a BODIPY 493/503-based flow cytometry screen, 6 negative and 3 positive regulators of fat content were identified. We selected one negative regulator of lipid content, C13C4.5, for future study. C13C4.5 was mainly expressed in the worm intestine. We found that this gene was important for maintaining the metabolism of lipid droplets. Biochemical results revealed that 50% of triacylglycerol (TAG) was lost in C13C4.5 knockout worms. Stimulated Raman scattering (SRS) signals in C13C4.5 mutants showed only 49.6% of the fat content in the proximal intestinal region and 86.3% in the distal intestinal region compared with wild type animals. The mean values of lipid droplet size and intensity in C13C4.5 knockout animals were found to be significantly decreased compared with those in wild type worms. The LMP-1-labeled membrane structures in worm intestines were also enlarged in C13C4.5 mutant animals. Finally, fertility defects were found in C13C4.5(ok2087) mutants. Taken together, these results indicate that C13C4.5 may regulate the fertility of C. elegans by changing the size and fat content of lipid droplets by interfering with lysosomal morphology and function.
Animals ; Biological Evolution ; Caenorhabditis elegans ; genetics ; growth & development ; metabolism ; Caenorhabditis elegans Proteins ; genetics ; metabolism ; Fertility ; Flow Cytometry ; Gene Knockout Techniques ; Humans ; Lipid Metabolism ; genetics ; Lysosomes ; genetics ; metabolism ; Membrane Proteins ; genetics ; Metabolic Networks and Pathways ; genetics ; Triglycerides ; metabolism

OBJECTIVETo examine the important roles of microRNAs (miRNAs) in regulating amphid structure and function, we performed a computational analysis for the genetic loci required for the sensory perception and their possibly corresponding miRNAs in C. elegans.

METHODSTotal 55 genetic loci required for the amphid structure and function were selected. Sequence alignment was combined with E value evaluation to investigate and identify the possible corresponding miRNAs.

RESULTSTotal 30 genes among the 55 genetic loci selected have their possible corresponding regulatory miRNA (s), and identified genes participate in the regulation of almost all aspects of amphid structure and function. In addition, our data suggest that both the amphid structure and the amphid functions might be regulated by a series of network signaling pathways. Moreover, the distribution of miRNAs along the 3' untranslated region (UTR) of these 30 genes exhibits different patterns.

CONCLUSIONWe present the possible miRNA-mediated signaling pathways involved in the regulation of chemosensation and thermosensation by controlling the corresponding sensory neuron and interneuron functions. Our work will be useful for better understanding of the miRNA-mediated control of the chemotaxis and thermotaxis in C. elegans.

Animals ; Caenorhabditis elegans ; embryology ; genetics ; Caenorhabditis elegans Proteins ; biosynthesis ; genetics ; Cilia ; genetics ; Computational Biology ; methods ; Gene Expression Regulation, Developmental ; genetics ; Genome ; genetics ; MicroRNAs ; genetics ; Models, Genetic ; Nervous System ; embryology ; metabolism ; Neurons, Afferent ; metabolism ; Sensation ; genetics ; Signal Transduction ; genetics

Animals integrate various environmental stimuli within the nervous system to generate proper behavioral responses. However, the underlying neural circuits and molecular mechanisms are largely unknown. The insulin-like signaling pathway is known to regulate dauer formation, fat metabolism, and longevity in Caenorhabditis elegans (C. Elegans). Here, we show that this highly conserved signaling pathway also functions in the integrative response to an olfactory diacetyl and a gustatory Cu(2+) stimuli. Worms of wild-type N2 Bristol displayed a strong avoidance to the Cu(2+) barrier in the migration pathway to the attractive diacetyl. Mutants of daf-2 (insulin receptor), daf-18 (PTEN lipid phosphatase), pdk-1 (phosphoinositide-dependent kinase), akt-1/-2 (Akt/PKB kinase) and sgk-1 (serum- and glucocorticoid-inducible kinase) show severe defects in the elusion from the Cu(2+). Mutations in DAF-16, a forkhead-type transcriptional factor, suppress the integrative defects of daf-2 and akt-1/-2 mutants. We further report that neither cGMP nor TGFβ pathways, two other dauer formation regulators, likely plays a role in the integrative learning. These results suggest that the insulin-like signaling pathway constitutes an essential component for sensory integration and decision-making behavior plasticity.
Animals ; Caenorhabditis elegans ; genetics ; physiology ; Caenorhabditis elegans Proteins ; genetics ; physiology ; Chemotaxis ; genetics ; physiology ; Copper ; physiology ; Cyclic GMP ; genetics ; physiology ; Diacetyl ; metabolism ; Insulin ; metabolism ; Longevity ; Signal Transduction ; Smell ; genetics ; physiology ; Taste ; genetics ; physiology ; Transforming Growth Factor beta ; genetics ; physiology