GLB-3: A resilient, cysteine-rich, membrane-tethered globin expressed in the reproductive and nervous system of Caenorhabditis elegans.

The popular genetic model organism Caenorhabditis elegans (C. elegans) encodes 34 globins, whereby the few that are well-characterized show divergent properties besides the typical oxygen carrier function. Here, we present a biophysical characterization and expression analysis of C. elegans globin-3 (GLB-3). GLB-3 is predicted to exist in two isoforms and is expressed in the reproductive and nervous system. Knockout of this globin causes a 99% reduction in fertility and reduced motility. Spectroscopic analysis reveals that GLB-3 exists as a bis-histidyl-ligated low-spin form in both the ferrous and ferric heme form. A function in binding of diatomic gases is excluded on the basis of the slow CO-binding kinetics. Unlike other globins, GLB-3 is also not capable of reacting with H2O2, H2S, and nitrite. Intriguingly, not only does GLB-3 contain a high number of cysteine residues, it is also highly stable under harsh conditions (pH = 2 and high concentrations of H2O2). The resilience diminishes when the N- and C-terminal extensions are removed. Redox potentiometric measurements reveal a slightly positive redox potential (+8 ± 19 mV vs. SHE), suggesting that the heme iron may be able to oxidize cysteines. Electron paramagnetic resonance shows that formation of an intramolecular disulphide bridge, involving Cys70, affects the heme-pocket region. The results suggest an involvement of the globin in (cysteine) redox chemistry.

Identification of healthspan-promoting genes in Caenorhabditis elegans based on a human GWAS study.

To find drivers of healthy ageing, a genome-wide association study (GWAS) was performed in healthy and unhealthy older individuals. Healthy individuals were defined as free from cardiovascular disease, stroke, heart failure, major adverse cardiovascular event, diabetes, dementia, cancer, chronic obstructive pulmonary disease (COPD), asthma, rheumatism, Crohn's disease, malabsorption or kidney disease. Six single nucleotide polymorphisms (SNPs) with unknown function associated with ten human genes were identified as candidate healthspan markers. Thirteen homologous or closely related genes were selected in the model organism C. elegans for evaluating healthspan after targeted RNAi-mediated knockdown using pathogen resistance, muscle integrity, chemotaxis index and the activity of known longevity and stress response pathways as healthspan reporters. In addition, lifespan was monitored in the RNAi-treated nematodes. RNAi knockdown of yap-1, wwp-1, paxt-1 and several acdh genes resulted in heterogeneous phenotypes regarding muscle integrity, pathogen resistance, chemotactic behaviour, and lifespan. Based on these observations, we hypothesize that their human homologues WWC2, CDKN2AIP and ACADS may play a role in health maintenance in the elderly.

Prediction of biological age by morphological staging of sarcopenia in Caenorhabditis elegans.

Sarcopenia encompasses a progressive decline in muscle quantity and quality. Given its close association with ageing, it may represent a valuable healthspan marker. The commonalities with human muscle structure and facile visualization possibilities make Caenorhabditis elegans an attractive model for studying the relationship between sarcopenia and healthspan. However, classical visual assessment of muscle architecture is subjective and has low throughput. To resolve this, we have developed an image analysis pipeline for the quantification of muscle integrity in confocal microscopy images from a cohort of ageing myosin::GFP reporter worms. We extracted a variety of morphological descriptors and found a subset to scale linearly with age. This allowed establishing a linear model that predicts biological age from a morphological muscle signature. To validate the model, we evaluated muscle architecture in long-lived worms that are known to experience delayed sarcopenia by targeted knockdown of the daf-2 gene. We conclude that quantitative microscopy allows for staging sarcopenia in C. elegans and may foster the development of image-based screens in this model organism to identify modulators that mitigate age-related muscle frailty and thus improve healthspan.