Beardsley, Simon et al. (2021) International Worm Meeting "Loss of adaptor protein complexes bypasses mrp-5 deficiency and restores heme deficit in Caenorhabditis elegans"

Dutt, Sohini, Beardsley, Simon, Yuan, Xiaojing, Hamza, Iqbal, Smith, Harold, Krause, Mike

[International Worm Meeting, 2021]

Metazoans require heme as an essential cofactor and synthesize it in the mitochondria. Caenorhabditis elegans and related helminths are an exception as they are unable to synthesize heme de novo but instead acquire heme from dietary sources for their growth and development. Heme is imported from the apical surface of the intestine by HRG-1-related importers and exported by MRP-5/ABCC5 from the basolateral surface. Loss of mrp-5 causes heme deficiency in extra-intestinal tissues and embryonic lethality, which can be suppressed by dietary heme supplementation raising the possibility that heme export in the absence of mrp-5 could be mediated by alternate pathways. Here we show, by performing a forward genetic screen in an mrp-5 null mutant, that loss-of-function in vesicular transport adaptor protein complexes fully restores heme homeostasis and normal growth and development in mrp-5 mutants. We screened over 160,000 haploid genomes and identified thirty-two bypass suppressors of mrp-5(ok2067). Deep-sequencing and variant analyses revealed three suppressors, each containing mutations in different adaptor protein 3 (AP3) complex subunits. RNAi depletion of individual subunits for either AP3 or AP2 phenocopies the mrp-5 bypass suppressors. Remarkably, loss of AP3 subunits in the mrp-5 mutants reversed intestinal accumulation of fluorescent heme analog; resistance to gallium protoporphyrin IX toxicity; extra-intestinal heme deficiencies; and neuronal abnormalities including glial migration and synaptogenesis. Taken together, our results show that cargo transport by vesicular adaptor proteins play a previously unanticipated role in systemic heme homeostasis and that multiple routes govern intra- and intercellular heme trafficking in metazoans.

Yu, Simon et al. (2011) International Worm Meeting "Deciphering mechanisms by which EGF signaling protects against the aging process."

Driscoll, Monica, Iwasa, Hiroaki, Xue, Jian, Yu, Simon

[International Worm Meeting, 2011]

We previously reported that EGF signaling confers a positive effect on multiple aging phenotypes, acting through the downstream branch of the EGF pathway involving PLCgand the IP3 receptor (Iwasa et al., 2010).To better understand operative mechanisms, we are analyzing mechanistic details of this healthspan pathway, looking at ligand requirements, testing other signaling components, and screening for downstream effectors. EGF ligand acts throughout development to influence healthy aging. lin-3 encodes the C. elegans EGF ligand, which can be expressed as multiple isoforms due to alternative splicing (Dutt et al., 2004, Van Buskirk et al., 2007). Little attention has been given to adult expression pattern. We followed a lin-3::gfp fusion in adults and conclude that lin-3 is expressed late into adult life in the pharynx.We tested for effects of over-expression of the intact gene and individual isoforms to address whether any one might promote locomotory healthspan extension and lifespan. We identify two of the individual LIN-3 isoforms that show anti-aging effects. Another question of interest is: when does LIN-3/EGF act to influence aging quality? We used RNAi approaches to show that LIN-3 is required both at larval stages and adult stages to influence maintained swimming performance and lifespan late in life. Implications of this finding are that: 1) EGF promotes maintenance throughout life, 2) effects of EGF signaling in development can have an impact later in life, and 3) EGF acts during adulthood to promote healthy aging. Another unresolved question is in what tissues does EGF signaling act to promote healthy aging. We expressed the let-23(gf) EGF receptor in specific tissues and asked whether age-associated mobility decline and lifespan are beneficially affected. We found ectopic expression of let-23(gf) in the intestine, neuron, and muscle can promote healthier aging phenotypes. Our previous data show that activated EGFR acts via increased IP3 activity through the IP3 receptor ER calcium release channel to confer healthspan benefits. We found that other known factors that modulate EGFR (ark-1) and control IP3 signaling and ER calcium release, including kinase (lfe-2) and phosphatase (ipp-5) can influence healthspan, consistent with the involvement of the core pathway in healthy aging. Searches for downstream factors that might mediate calcium signaling coming from EGF activation. Our data implicate appropriate/maintained calcium homeostasis in healthy aging. Though many factors could execute ER release and downstream calcium signaling, we identified one possible calcium-sensitive transcription factor downstream of IP3R in the EGF pathway that can regulate the effects on locomotory healthspan.