A Caenorhabditis elegans model for ether lipid biosynthesis and function

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Shi Z, Tarazona P, Brock TJ, Browse J, Feussner I, and Watts JL. (2016) A Caenorhabditis elegans model for ether lipid biosynthesis and function. Journal of Lipid Research, 57.

Although being studied for nearly 100 years, the roles of plasmalogens, a phospholipid that contains a vinyl-ether bond at the sn-1 position, are still not fully known. In humans plasmalogens have been found in many body systems including nervous, immune, and cardiovascular systems. The diversity of mechanisms they are thought to be involved in and locations found in the body aid in keeping their functions elusive. Evolutionary conservation has allowed for simpler organisms to be utilized in determining the mechanisms behind these effects.

In this study by Shi and colleagues, a Caenorhabditis elegans (C. elegans) roundworm model was used to investigate the role of plasmalogens in survival and stress-response. The authors created several plasmalogen-deficient worm strains using RNA interference to knock down the expression of several plasmalogen biosynthetic genes including fatty acyl-CoA reductase (FAR-1; FARD-1), glyceronephosphate O-acyltransferase (GNPAT; ACL-7), or alkylglycerone phosphate synthetase (AGPS; ADS-1). Using a liquid chromatography tandem mass spectrometry (LC-MS/MS) assay, levels of different plasmalogen species were shown to vary within the wild-type N2 worms, while a near complete loss of plasmalogens was confirmed within the mutant strains. 

To characterize the effects of impaired plasmalogen synthesis the three mutant strains were first compared to wild-type worms with a battery of functional assays including cold tolerance, brood size, and lifespan and can be seen in the figure below. Wild-type worms grown at 10°C showed a 60% survival rate, while the acl-7 and ads-1 worms had survival rates of approximately 20% and fard-1 was near 5% (A). Wild-type C. elegans are able to survive at colder temperatures because they are capable of sensing light and pheromones in their environment and making the appropriate biochemical alterations in response. For this to be effective, a signal must be passed from the neurons that sense the stimuli, however plasmalogen deficiencies are known to negatively affect vesicular fusion, and could be impeding this process. Brood size was also reduced from an average of 300 progeny in wild-type worms to 200 in acl-7, and further to 150 in fard-1 and ads-1 worms (B). Of most relevance, the overall lifespans compared to wild-type of fard-1, ads-1, and acl-7 mutants were reduced by 29.0%, 29.6%, and 30.1%, respectively (C).

Second, the plasmalogen-deficient worms were tested for their sensitivity to various stress inducers. Treatment with tunicamycin, a chemical that causes endoplasmic reticulum (ER) stress, resulted in no significant increase in sensitivity, suggesting that ER stress responses are not plasmalogen-mediated (F). However, plasmalogens are known antioxidants and provide protection against reactive oxygen species (ROS). To test this, the wild-type and mutant strains were treated with high concentrations of paraquat, a chemical that generates ROS, which resulted in 50% death in the wild-type worms by 6 hours (D). Survival was further impaired in the mutant strains, where 80-90% died within the same amount of time. Tert-butyl peroxide, another ROS inducing agent, resulted in the death of wild-type worms within 8 hours, and all plasmalogen-deficient worms after only 6 hours of treatment (E). These results would suggest that the plasmalogen deficiency does negatively affect the worm’s ability to protect itself against ROS, making the worms an appropriate model to further analyze the antioxidant properties of plasmalogens.

Taken together, Shi et al have provided novel plasmalogen-deficient worm strains for future work to be completed in. As plasmalogen deficiencies cause a more severe phenotype in mice, including cataracts, eye development defects, and male infertility, this study has displayed that C. elegans provide a more viable model for determining the mechanistic role plasmalogens play in a whole organism. In addition, they underscore the evolutionary conservation of plasmalogens between such different organisms as microscopic nematodes and humans, and the criticalness of plasmalogens in fundamental cellular processes, including stress response and longevity. Consistent with numerous other findings over the past decade or more, plasmalogen reduction compromises numerous cellular functions and provides further support that therapeutic plasmalogen augmentation might represent a novel approach for positively modulating these pathways.

Physiological consequences of ether lipid deficiency. A: Ether lipid-deficient mutants show reduced survival when grown at 15 C, and fail to survive at 10 C. Two-tailed t-tests were used to determine the significance between the mean cold survival of each mutant strain compared with wild-type at the same temperature. * P<0.05, P<0.001. B: Ether-lipid deficient mutants have a reduced number of viable progeny at 20 C. Graphs show the range and average number of progeny in eight to ten individuals of each genotype. The brood size of each mutant is significantly different from wild-type, ** P<0.0001 as determined by one-way ANOVA and Dunnett’s multiple comparison test. C: Lifespan of ether-lipid deficient at 20 C is shorter than wild-type, P<0.0001 as determined by log-rank (Cox-Mantel) test. D, E: Ether-lipid deficient mutants show reduced survival in the presence of acute oxidative stress. For both oxidants, P<0.001 as determined by log-rank (Cox-Mantel) test. F: Ether lipid-deficient mutants are not more sensitive to ER stress than wild-type.