C. elegans Germline-Deficient Mutants Respond to Pathogen Infection Using Shared and Distinct Mechanisms

Reproduction extracts a cost in resources that organisms are then unable to utilize to deal with a multitude of environmental stressors. In the nematode C. elegans, development of the germline shortens the lifespan of the animal and increases its susceptibility to microbial pathogens. Prior studies have demonstrated germline-deficient nematodes to have increased resistance to Gram negative bacteria. We show that germline-deficient strains display increased resistance across a broad range of pathogens including Gram positive and Gram negative bacteria, and the fungal pathogen Cryptococcus neoformans. Furthermore, we show that the FOXO transcription factor DAF-16, which regulates longevity and immunity in C. elegans, appears to be crucial for maintaining longevity in both wild-type and germline-deficient backgrounds. Our studies indicate that germline-deficient mutants glp-1 and glp-4 respond to pathogen infection using common and different mechanisms that involve the activation of DAF-16

A polymorphic variant of the insulin-like growth factor 1 (IGF-1) receptor correlates with male longevity in the Italian population: a genetic study and evaluation of circulating IGF-1 from the "Treviso Longeva (TRELONG)" study

<p>Abstract</p> <p>Background</p> <p>An attenuation of the insulin-like growth factor 1 (IGF-1) signaling has been associated with elongation of the lifespan in simple metazoan organisms and in rodents. In humans, IGF-1 level has an age-related modulation with a lower concentration in the elderly, depending on hormonal and genetic factors affecting the IGF-1 receptor gene (<it>IGF-1R</it>).</p> <p>Methods</p> <p>In an elderly population from North-eastern Italy (<it>n </it>= 668 subjects, age range 70–106 years) we investigated the <it>IGF-1R </it>polymorphism G3174A (<it>rs2229765</it>) and the plasma concentration of free IGF-1. Frequency distributions were compared using χ²-test "Goodness of Fit" test, and means were compared by one-way analysis of variance (ANOVA); multiple regression analysis was performed using JMP7 for SAS software (SAS Institute, USA). The limit of significance for genetic and biochemical comparison was set at α = 0.05.</p> <p>Results</p> <p>Males showed an age-related increase in the A-allele of <it>rs2229765 </it>and a change in the plasma level of IGF-1, which dropped significantly after 85 years of age (85+ group). In the male 85+ group, A/A homozygous subjects had the lowest plasma IGF-1 level. We found no clear correlation between <it>rs2229765 </it>genotype and IGF-1 in the females.</p> <p>Conclusion</p> <p>These findings confirm the importance of the <it>rs2229765 </it>minor allele as a genetic predisposing factor for longevity in Italy where a sex-specific pattern for IGF-1 attenuation with ageing was found.</p

Lensfree Fluorescent On-Chip Imaging of Transgenic Caenorhabditis elegans Over an Ultra-Wide Field-of-View

We demonstrate lensfree on-chip fluorescent imaging of transgenic Caenorhabditis elegans (C. elegans) over an ultra-wide field-of-view (FOV) of e.g., >2–8 cm2 with a spatial resolution of ∼10µm. This is the first time that a lensfree on-chip platform has successfully imaged fluorescent C. elegans samples. In our wide-field lensfree imaging platform, the transgenic samples are excited using a prism interface from the side, where the pump light is rejected through total internal reflection occurring at the bottom facet of the substrate. The emitted fluorescent signal from C. elegans samples is then recorded on a large area opto-electronic sensor-array over an FOV of e.g., >2–8 cm2, without the use of any lenses, thin-film interference filters or mechanical scanners. Because fluorescent emission rapidly diverges, such lensfree fluorescent images recorded on a chip look blurred due to broad point-spread-function of our platform. To combat this resolution challenge, we use a compressive sampling algorithm to uniquely decode the recorded lensfree fluorescent patterns into higher resolution images, demonstrating ∼10 µm resolution. We tested the efficacy of this compressive decoding approach with different types of opto-electronic sensors to achieve a similar resolution level, independent of the imaging chip. We further demonstrate that this wide FOV lensfree fluorescent imaging platform can also perform sequential bright-field imaging of the same samples using partially-coherent lensfree digital in-line holography that is coupled from the top facet of the same prism used in fluorescent excitation. This unique combination permits ultra-wide field dual-mode imaging of C. elegans on a chip which could especially provide a useful tool for high-throughput screening applications in biomedical research

Lifespan Extension by Preserving Proliferative Homeostasis in Drosophila

Regenerative processes are critical to maintain tissue homeostasis in high-turnover tissues. At the same time, proliferation of stem and progenitor cells has to be carefully controlled to prevent hyper-proliferative diseases. Mechanisms that ensure this balance, thus promoting proliferative homeostasis, are expected to be critical for longevity in metazoans. The intestinal epithelium of Drosophila provides an accessible model in which to test this prediction. In aging flies, the intestinal epithelium degenerates due to over-proliferation of intestinal stem cells (ISCs) and mis-differentiation of ISC daughter cells, resulting in intestinal dysplasia. Here we show that conditions that impair tissue renewal lead to lifespan shortening, whereas genetic manipulations that improve proliferative homeostasis extend lifespan. These include reduced Insulin/IGF or Jun-N-terminal Kinase (JNK) signaling activities, as well as over-expression of stress-protective genes in somatic stem cell lineages. Interestingly, proliferative activity in aging intestinal epithelia correlates with longevity over a range of genotypes, with maximal lifespan when intestinal proliferation is reduced but not completely inhibited. Our results highlight the importance of the balance between regenerative processes and strategies to prevent hyperproliferative disorders and demonstrate that promoting proliferative homeostasis in aging metazoans is a viable strategy to extend lifespan

RNAi Screen of DAF-16/FOXO Target Genes in C. elegans Links Pathogenesis and Dauer Formation

The DAF-16/FOXO transcription factor is the major downstream output of the insulin/IGF1R signaling pathway controlling C. elegans dauer larva development and aging. To identify novel downstream genes affecting dauer formation, we used RNAi to screen candidate genes previously identified to be regulated by DAF-16. We used a sensitized genetic background [eri-1(mg366); sdf-9(m708)], which enhances both RNAi efficiency and constitutive dauer formation (Daf-c). Among 513 RNAi clones screened, 21 displayed a synthetic Daf-c (SynDaf) phenotype with sdf-9. One of these genes, srh-100, was previously identified to be SynDaf, but twenty have not previously been associated with dauer formation. Two of the latter genes, lys-1 and cpr-1, are known to participate in innate immunity and six more are predicted to do so, suggesting that the immune response may contribute to the dauer decision. Indeed, we show that two of these genes, lys-1 and clc-1, are required for normal resistance to Staphylococcus aureus. clc-1 is predicted to function in epithelial cohesion. Dauer formation exhibited by daf-8(m85), sdf-9(m708), and the wild-type N2 (at 27°C) were all enhanced by exposure to pathogenic bacteria, while not enhanced in a daf-22(m130) background. We conclude that knockdown of the genes required for proper pathogen resistance increases pathogenic infection, leading to increased dauer formation in our screen. We propose that dauer larva formation is a behavioral response to pathogens mediated by increased dauer pheromone production

DAF-21/Hsp90 is required for C. elegans longevity by ensuring DAF-16/FOXO isoform A function

The FOXO transcription factor family is a conserved regulator of longevity and the downstream target of insulin/insulin-like signaling. In Caenorhabditis elegans, the FOXO ortholog DAF-16A and D/F isoforms extend lifespan in daf-2 insulin-like receptor mutants. Here we identify the DAF-21/Hsp90 chaperone as a longevity regulator. We find that reducing DAF-21 capacity by daf-21(RNAi) initiated either at the beginning or at the end of larval development shortens wild-type lifespan. daf-21 knockdown employed from the beginning of larval development also decreases longevity of daf-2 mutant and daf-2 silenced nematodes. daf-16 loss-of-function mitigates the lifespan shortening effect of daf-21 silencing. We demonstrate that DAF-21 specifically promotes daf-2 and heat-shock induced nuclear translocation of DAF-16A as well as the induction of DAF-16A-specific mRNAs, without affecting DAF-16D/F localization and transcriptional function. DAF-21 is dispensable for the stability and nuclear import of DAF-16A, excluding a chaperone-client interaction and suggesting that DAF-21 regulates DAF-16A activation upstream of its cellular traffic. Finally, we show a selective requirement for DAF-21 to extend lifespan of DAF-16A, but not DAF-16D/F, transgenic daf-2 mutant strains. Our findings indicate a spatiotemporal determination of multiple DAF-21 roles in fertility, development and longevity and reveal an isoform-specific regulation of DAF-16 activity. © 2018, The Author(s)

Metalloproteins and phytochelatin synthase may confer protection against zinc oxide nanoparticle induced toxicity in Caenorhabditis elegans

Zinc oxide nanoparticles (ZnONPs) are used in large quantities by the cosmetic, food and textile industries. Here we exposed Caenorhabditis elegans wild-type and a metal sensitive triple knockout mutant (mtl-1;mtl-2;pcs-1) to ZnONPs (0-50mg/L) to study strain and exposure specific effects on transcription, reactive oxygen species generation, the biomolecular phenotype (measured by Raman microspectroscopy) and key endpoints of the nematode life cycle (growth, reproduction and lifespan). A significant dissolution effect was observed, where dissolved ZnO constituted over 50% of total Zn within a two day exposure to the test medium, suggesting that the nominal exposure to pure ZnONPs represents in vivo, at best, a mixture exposure of ionic zinc and nanoparticles. Nevertheless, the analyses provided evidence that the metallothioneins (mtl-1 and mtl-2), the phytochelatin synthase (pcs-1) and an apoptotic marker (cep-1) were transcriptionally activated. In addition, the DCFH-DA assay provided in vitro evidence of the oxidative potential of ZnONPs in the metal exposure sensitive triple mutant. Raman spectroscopy highlighted that the biomolecular phenotype changes significantly in the mtl-1;mtl-2;pcs-1 triple knockout worm upon ZnONP exposure, suggesting that these metalloproteins are instrumental in the protection against cytotoxic damage. Finally, ZnONP exposure was shown to decrease growth and development, reproductive capacity and lifespan, effects which were amplified in the triple knockout. By combining diverse toxicological strategies, we identified that individuals (genotypes) housing mutations in key metalloproteins and phytochelatin synthase are more susceptible to ZnONP exposure, which underlines their importance to minimize ZnONP induced toxicity