Balasubramaniam, Meenakshisundaram et al. (2013) International Worm Meeting "Role of amino-acid pool size in aging-related muscle atrophy."

Ayyadevara, Srinivas, Reis, Robert J Shmookler, Alla, Ramani, Balasubramaniam, Meenakshisundaram

[International Worm Meeting, 2013]

Skeletal muscle mass declines to varying degrees with age and this is a significant contributor to frailty, loss of mobility and increased mortality. Reduced muscle mass can result from a decrease in muscle synthesis and/or an increase in degradation, as a consequence of trauma, aging and many age-onset disorders. A previous NMR study reported that a long-lived mutant strain has elevated pool levels of several amino acids, in particular the branched-chain amino acids leucine, isoleucine, valine, phenylalanine, tyrosine and tryptophan. To learn whether specific amino acid pools are generally associated with improved muscle retention or lifespan across diverse longevity mutants, we have determined the free aminoacid levels in a panel of C.elegans longevity mutants.In our initial assay we found increased pools of several branched-chain amino acids in the long-lived mutant strains. Remarkably, dietary supplementation of these aminoacids reduced age-dependent sarcopenia and moderately extended life span for a wild-type strain. Based on these results in a model system, we predict that prolonged amino acid supplementation may enhance muscle mass, strength and function in older persons by increasing muscle growth and quality.

Suri, Pooja et al. (2015) International Worm Meeting "Role of aggregation-prone proteins in age-related disease."

Suri, Pooja, Alla, Ramani, Reis, Robert Shmookler, Balasubramaniam, Meenakshisundaram, Ayyadevara, Srinivas

[International Worm Meeting, 2015]

Protein aggregation is caused by a multitude of factors; there could be specific environmental causes like oxidative stress, or a genetic predisposition to aggregation. Protein aggregation increases with aging, as the ability to rectify or clear misfolded proteins is reduced. This is partly due to age-dependent deterioration in the ubiquitin/proteasomal system or other unfolded protein response systems. Aggregation will be enhanced when a misfolded, dysfunctional protein does not undergo degradation via normal pathways like autophagy/lysosome or ubiquitin proteasome-mediated degradation pathways. By mass spectrometry, we identified several proteins in the aggregates of a C. elegans model of Huntington's Disease. The effects on protein aggregation, of RNAi-mediated knockdown of some of these proteins, were tested. Some of the gene knock downs by RNAi altered the extent of aggregation in C. elegans, suggesting that these could act as seed proteins and enhance the progression of aggregates. These can be useful targets to disrupt the aggregates and could be exploited for therapeutic interventions in neurological diseases.

ayyadevara, srinivas et al. (2019) International Worm Meeting "A novel microtubule-binding drug attenuates and reverses protein aggregation in animal models of Alzheimer's disease."

Kakraba, Samuel, Alla, Ramani, Penthala, Narsimha Reddy, Griffin, Sue, Ganne, Akshatha, Barger, Steven, Ayyadevara, Srinivas, Liu , Ling, Shmookler Reis, Robert, Balasubramaniam, Meenakshisundaram, Bommagani, Shoban Babu, Crooks, Peter

[International Worm Meeting, 2019]

Age-progressive neurodegenerative pathologies, including Alzheimer's disease, are distinguished and diagnosed by disease-specific components of intra- or extra-cellular aggregates. Increasing evidence suggests that neuroinflammation promotes protein aggregation, and is involved in the etiology of neurological diseases. We synthesized and tested analogues of the naturally occurring tubulin-binding compound, combretastatin A-4. One such analogue, PNR502, markedly reduced the quantity of Alzheimer-associated amyloid aggregates in the BRI-A?1-42 mouse model of Alzheimer's disease, while blunting the ability of the pro-inflammatory cytokine IL-1? to raise levels of amyloid plaque and its protein precursors in a neuronal cell-culture model. In transgenic C. elegans strains that express human A?1-42 in muscle or neurons, PNR502 rescued A?-induced disruption of motility (3.8-fold, p<0.0001) or chemotaxis (1.8-fold, p<0.05), respectively. Moreover, in C. elegans with neuronal expression of A?1-42, a single day of PNR502 exposure reverses the chemotaxis deficit by 54% (p<0.01), actually exceeding the protection from longer exposure. Moreover, continuous PNR502 treatment also extends nematode lifespan 23% (p?0.001). Given that PNR502 can slow, prevent, or reverse Alzheimer-like protein aggregation in human-cell-culture and animal models, and that its principal predicted and observed binding targets are proteins previously implicated in Alzheimer's, we propose that PNR502 has therapeutic potential to inhibit cerebral A?1-42 aggregation and prevent or reverse neurodegeneration.

Chun-Chieh Chen et al. (2002) East Coast Worm Meeting "Biochemical and genetic analysis of RNA interference in C. elegans"

Chun-Chieh Chen, Darryl Conte, Hiroaki Tabara, Craig Mello

[East Coast Worm Meeting, 2002]

RNAi is a post-transcriptional gene-silencing phenomenon induced by double-stranded RNAs. During RNAi, small-interfering RNAs (siRNAs) are formed and are thought to guide a nuclease complex in the destruction of target mRNAs. Interestingly, in C. elegans, the presence of a complementary target gene is required for siRNA accumulation and only the antisense siRNA strand is detectable by Northern analysis (Alla Grishok unpublished) This asymmetric accumulation of siRNA differs from results observed in Drosophila and plants and suggests that RNAi in C. elegans may include a surveillance step that ensures that only "useful" siRNAs are retained or amplified. Interestingly, RDE-4 has recently been shown to interact with RDE-1, DCR-1 and the trigger dsRNA, presumably forming a complex that functions to detect the presence of target mRNA (H. Tabara et al.). To further understand the mechanism of the target-dependent asymmetric accumulation of siRNAs, we performed northern analysis of wild-type and RNAi-deficient mutant worms exposed to pos-1 dsRNAs by feeding. We found that no siRNAs accumulates in dcr-1, rde-1 or rde-4 mutants. The absence of siRNA in rde-1 and rde-4 mutant backgrounds is consistent with the finding that these two proteins are required for the initiation of RNAi. Our findings also demonstrate for the first time that DCR-1 activity is indeed required for siRNA production in vivo. Consistent with a block in dsRNA processing we found that long-trigger dsRNA molecules accumulate in the dcr-1 mutants. Further biochemical tests are planned to examine the production of siRNAs by the DCR-1 complex. In addition, a new genetic screen will be carried out to identify additional rde mutants.

Bernstein, Max et al. (2013) International Worm Meeting "The hunt for quantitative trait nucleotides: a near-isogenic line based approach in C. elegans."

Rockman, Matthew, Bernstein, Max

[International Worm Meeting, 2013]

The individual nucleotides responsible for phenotypic diversity have been very difficult to isolate. The majority of known causal variants that affect traits in wild populations of a species occur within protein-coding regions. These variants are typically of very large effect and may not be typical of the sorts of alleles that drive evolution. There are many confounding effects that can explain the relative lack of causal variants, or quantitative trait nucleotides (QTNs), particularly alleles of small effect, epistasis, and gene-environment interactions. One way to elucidate quantitative trait loci (QTLs) is through the creation of near-isogenic lines (NILs), where the majority of the genome (>95%) is of one genomic background. NILs have previously been created for the nematode C. elegans and identified QTLs across the genome for several phenotypes. C. elegans is a naturally inbred species, which makes it ideal for isolating QTNs. Starting with a NIL that harbors a ~1.5 Mb segment on Chromosome X originating from the Hawaiian isolate CB4856, we have created a series of more than 1000 sub-NILs that break up this segment into smaller regions by recombination. These sub-NILs will be genotyped at 278 SNPs within the 1.5 Mb segment. We are currently piloting high-throughput fitness assays similar to previous work by Elvin et al (2011) and Ramani et al (2012). Additionally, we plan to do gene expression analysis on a subset of the sub-NIL panel to characterize expression QTNs (eQTNs). QTL mapping on simulated sub-NIL datasets shows that for a single QTL model, we can accurately map small effect (h2<0.1) causal variants. Due to the linkage disequilibrium pattern in the sub-NIL panel, traditional QTL mapping methods are not applicable. We show that pairwise comparisons among groups of similar sub-NIL genotypes provide a systematic method for identifying QTLs. We have also modeled more complex QTL scenarios, including multiple QTLs and epistatic interactions. These simulations have informed us of our potential statistical power for future QTL mapping experiments.

Nabih, Amena et al. (2015) International Worm Meeting "Exploring roles for C. elegans Argonaute/small RNA pathways in splicing."

Tu, Shikui, Claycomb, Julie M., Weng, Zhiping, Wedeles, Christopher J., Nabih, Amena

[International Worm Meeting, 2015]

Our understanding of endogenous small RNA pathways has grown dramatically over the past twenty years to encompass a host of gene regulatory activities, ranging from mRNA decay and translational repression in the cytoplasm, to transcriptional modulation in the nucleus. In these pathways, Argonautes (AGOs) are guided to target transcripts by small RNA binding partners via sequence complementarity. Upon recruitment to their targets, Argonautes in turn engage other regulatory proteins to effectively execute various gene regulatory outcomes. Although many aspects of small RNA mediated gene regulation have been studied at depth, one that has yet to be explored thoroughly is the role of Argonaute/small RNA pathways in splicing. Several recent studies have implicated human Argonaute/small RNA pathways in influencing alternative splicing (Allo et al., 2014, Ameyar-Zazoua et al., 2012). Conversely, stalled spliceosomes have been shown to trigger small RNA mediated genome defense in fungi (Dumesic et al. 2013). These tantalizing initial results open the possibility that splicing and small RNA pathways could be intricately linked to provide combinatorial regulation of gene expression and/or genome defense over a broad range of species.With 26 Argonautes and four types of endogenous small RNAs, C. elegans possesses a rich tapestry of small RNA mediated gene regulatory activity. In C. elegans, thus far one report has implicated small RNA pathways (CSR-1/22G-RNA and PRG-1/piRNA) in the alternative splicing of a single transcript, tor (target of rapamycin) (Barberan-Soler et al., 2014). CSR-1 is an essential Argonaute that plays a key role in positively regulating the majority of germline transcripts at the transcriptional level (Wedeles, Wu and Claycomb, 2013; Seth et al., 2013; Conine et al., 2013; Cecere et al., 2014). Two additional studies have revealed changes in alternative splicing patterns for a number of germline genes that are also the targets of the CSR-1 small RNA pathway (Ramani et al., 2010; Ortiz et al., 2014), opening the possibility that the CSR-1 small RNA pathway could play important roles in splicing germline transcripts. A role for CSR-1 in splicing is further supported by our preliminary data, which show a physical interaction with a conserved splicing factor. Ongoing transcriptome and phenotypic analysis of various strains in which csr-1 or the splicing factor are lost will shed led light on the impact of these factors on splicing across the genome, and may provide key insights into the role of small RNA pathways in splicing during C. elegans development. .