Chavez, Ivan et al. (2021) International Worm Meeting "Skin-penetrating nematodes exhibit life-stage-specific interactions with host-associated and environmental bacteria"

Chavez, Ivan, Bryant, Astra, Assie, Adrien, Samuel, Buck, Hallem, Elissa, Brown, Taylor

[International Worm Meeting, 2021]

Skin-penetrating nematodes of the genus Strongyloides infect over 600 million people, posing a major global health burden. Their life cycle includes both a parasitic and free-living generation. During the parasitic generation, infective third-stage larvae (iL3s) actively engage in host seeking. During the free-living generation, the nematodes develop and reproduce on host feces. At different points of their life cycle, Strongyloides species encounter bacteria from various ecological niches. However, the microbial interactions between Strongyloides and bacteria remain uncharacterized. We first investigated the microbiome of the human parasite Strongyloides stercoralis using 16S-based amplicon sequencing. We found that S. stercoralis free-living adults have a distinct microbiome, suggesting that they selectively associate with specific fecal bacteria. We then investigated the behavioral responses of S. stercoralis and the closely related rat parasite Strongyloides ratti to an ecologically diverse panel of bacteria. We found that S. stercoralis and S. ratti showed similar responses to bacteria. The responses of both nematodes to bacteria varied dramatically across life stages: free-living adults were strongly attracted to most of the bacteria tested, while iL3s were attracted specifically to soil bacteria. The behavioral responses to bacteria were dynamic, consisting of distinct short- and long-term behaviors. Finally, a comparison of the growth and reproduction of S. stercoralis free-living adults on different bacteria revealed that the bacterium Proteus mirabilis inhibits S. stercoralis egg hatching, greatly decreasing parasite viability. Our results identify bacteria that serve as key sensory cues for directing movement, as well as bacteria that decrease the parasite's reproductive fitness.

Rodrigues, Pedro Reis et al. (2009) International Worm Meeting "A variety of age-dependant aggregating proteins determine lifespan."

Peters, Theodore, Gibson, Brad, Lithgow, Gordon, Hughes, Robert, Alavez, Silvestre, Rodrigues, Pedro Reis, Czerwieniec, Gregg

[International Worm Meeting, 2009]

Protein aggregation has for long been hypothesised as a determinant of lifespan. Briefly, normal cellular activity may give rise to damaged proteins causing them to become insoluble, missfold and aggregate. To test this hypothesis we adapted a protocol in order to extract insoluble proteins from synchronously aging populations of C. elegans. Proteins were separated based on their aqueous and detergent solubility and the insoluble fraction was resolubilized in 70%; formic acid. Insoluble proteins were chemically labelled, identified and quantified by liquid chromatography coupled with mass spectrometry (LC- ESI-MS/MS). We identified a range of proteins with roles in various cellular processes and possibly from a range of cellular compartments. 27%; of the proteins identified as forming aggregates have previously been shown to be important in keeping low levels of polyglutamine aggregation1. This suggests that reduced soluble levels of these proteins caused by age-related aggregation may cause increased risk of polyglutamine aggregation. We then considered whether proteins that appear to form aggregates during normal aging influenced lifespan. To test this notion we, reduced their expression in adult animals (from 4 days old) by RNA interference (RNAi). 34%; of the RNAi treatments were found to significantly extend mean lifespan in C. elegans suggesting that a variety of age-dependant aggregating proteins determine lifespan. Among the insoluble proteins, DAF-21, an ortholog of the mammalian HSP-90, showed age-dependant aggregation and is being used as a marker to study the role of several molecular pathways in protein aggregation. Taken together our results suggest that protein aggregation may play a common and key role in aging and age-related disease. 1 - Nollen, E., Garcia, S., Haaften, G., Kim, S., Chavez, A., Morimoto, R., Plasterk, R., Genome-wide RNA interference screen identified previously undescribed regulators of polyglutamine aggregation. Proc Natl. Acad. Sci., 101, 6403-6408, 2004.

Ellen Nollen et al. (2006) European Worm Meeting "C. elegans Models of Protein Misfolding Diseases"

Ellen Nollen, Tjakko van Ham, Ronald H. A. Plasterk.

[European Worm Meeting, 2006]

Ellen Nollen, Tjakko van Ham & Ronald H. A. Plasterk. Aggregation of misfolded proteins occurs in various age-related neurodegenerative disorders, including Parkinsons, Alzheimers, and Huntingtons disease. To understand how cells protect themselves against misfolded proteins, we search for genes that enhance or prevent protein aggregation. C. elegans strains expressing polyglutamine stretches fused to YFP with visible, age-dependend protein aggregation are used as a genetic model. Using a genome-wide RNAi screen, we have previously identified 186 genes that, when knocked down, cause premature protein aggregation. These genes include genes involved in protein synthesis, folding, degradation and RNA synthesis and processing. 1. Conversely, we performed a forward mutagenesis screen to identify genes that, when mutated, suppress age-dependent polyglutamine aggregation. For one suppressor mutant, in which aggregation is suppressed by more than 75%, we have now identified the responsible mutation. This mutation is a missense mutation in a gene encoding a protein of unknown function that is highly conserved between C. elegans and humans. Knock-down by RNAi of the same gene in wild-type worms yielded a similar reduction in aggregation, suggesting a loss-of-function mutation. We are currently further characterizing this mutant and the remaining suppressor mutants. In addition, to establish whether the genes we have identified are specific for polyglutamine aggregation or whether they comprise of a general protein homeostatic buffer, we have developed a worm model for aggregation of alpha synuclein, which occurs in Parkinson''s disease. Altogether our results will provide insight into cellular protection against misfolded proteins and yield targets for therapy against protein misfolding diseases.. 1Nollen E.A.A., Garcia S.M., van Haaften G., Kim S., Chavez A., Morimoto R.I., Plasterk R.H. (2004) Genome-wide RNA interference screen identifies previously undescribed regulators of polyglutamine aggregation. Proc. Natl. Acad. Sci. U.S.A. 101(17):6403-8.

T.J. van Ham et al. (2007) International Worm Meeting "A C. elegans model links novel modifiers of a-synuclein inclusion formation to aging."

K.L. Thijssen, E.A.A. Nollen, T.J. van Ham, R.H.A. Plasterk

[International Worm Meeting, 2007]

Accumulation and aggregation of <font face=symbol>a</font>-synuclein in the brain is characteristic of, and considered to be causatively related to, several age-related neurodegenerative diseases such as Parkinsons disease (PD). To study early stages of <font face=symbol>a</font>S accumulation and to identify modifier genes, we created transgenic C. elegans expressing the human <font face=symbol>a</font>S gene fused to YFP in the body wall muscle. These transgenic animals show localization of <font face=symbol>a</font>S-YFP to discrete foci during aging, which is not seen for YFP alone, indicating specificity to <font face=symbol>a</font>S. These foci are present as early as day two after hatching, and become more abundant during aging. By fluorescence recovery after photobleaching (FRAP) we determined that the foci contain mobile material until day 11. However, at old age also immobilized aggregated protein is present. The amount of <font face=symbol>a</font>S-foci increases to day 11 and decreases thereafter. Interestingly, this decrease in foci formation is associated with a relative increase in the amount of immobilized aggregates. We performed a genome-wide RNAi screen for modifiers of foci formation, and found 108 genes that repeatedly showed increased foci formation when silenced. Remarkably, none of these genes overlaps with genes found in a screen for polyglutamine aggregation we performed in C. elegans1. This might indicate that different mechanisms are involved in these diseases. To further characterize genes found in the screen, deletion strains of a selection of the genes representing different functional classes were crossed into the <font face=symbol>a</font>S-YFP strain. Deletion of a gene - involved in aging and encoding one of the most potent suppressors of foci-formation - resulted in a twofold increase in the amount of foci on day thirteen. We are currently further characterizing the deletion strains. In conclusion, we developed a new animal model that captures features of age-related PD. We used the model to dentify novel modifiers of <font face=symbol>a</font>S-YFP foci formation, of which we are further characterizing the effect of deletion of these genes on <font face=symbol>a</font>S-YFP foci formation. These results can provide insight into the early cellular mechanisms involved in <font face=symbol>a</font>S accumulation, and could thereby yield molecular targets for therapy. 1 Nollen E.A.A., Garcia S.M., van Haaften G., Kim S., Chavez A., Morimoto R.I., Plasterk R.H.A. (2004) Genome-wide RNA interference screen identifies previously undescribed regulators of polyglutamine aggregation. Proc. Natl. Acad. Sci. U.S.A. 101(17):6403-8.