Organisms modulate metabolic pathways dependent on life stage, nutrition, and environment, among other factors. CLK-1, an ortholog of human COQ7, is necessary for ubiquinone biosynthesis (Liu et al., 2017, PMID: 28404998). Mutation of
clk-1 results in many pleiotropic effects, including a slowing down of metabolic and rhythmic processes. The paralogous
fstr-1 and
fstr-2 genes were identified by the ability of their downregulation by RNAi to restore the speed of rhythmic behaviors that are slowed in
clk-1 mutants (Cristina et al., 2009, PMID: 19360127). FSTR-1/F57F4.3 and FSTR-2/F57F4.4 are paralogous proteins that are 96% identical and differ primarily by the absence of a small 63-aa region in FSTR-2. Both proteins are each >2000 amino acids long and contain many repeated ET domains of unknown function. We have previously found that strains in which gut specification is partially compromised ('hypomorphic gut specification' or HGS strains), animals accumulate extra lipids (Maduro et al., 2015, PMID: 25959238). Through RNA-seq analysis of intestine-enriched genes, we found that in wild-type strains,
fstr-2/F57F4.4 is upregulated, while in HGS strains,
fstr-1/F57F4.3 is upregulated. We hypothesized, therefore, that FSTR-1 and FSTR-2 play related but differentiable roles in regulating metabolism. Consistent with a focus in the intestine, wrmScarlet reporters (transcriptional and translational) are enriched in the gut, with primarily cytoplasmic localization. To further characterize the roles of
fstr-1 and
fstr-2 in metabolism, we generated single and double knockout alleles by CRISPR/Cas9-mediated mutagenesis. Both
fstr-1 fstr-2 double mutants and
fstr-2 single mutants appear to contain more gut granules and have a different distribution of lipids than controls, suggesting that FSTR-2 has a role in regulation of normal metabolism in the intestine. While we find no evidence of a changed life span in
fstr-1 fstr-2 double mutants, these animals exhibit increased susceptibility to toxin-mediated killing by Pseudomonas aeruginosa. Others have reported upregulation of
fstr-2::GFP during pathogen infection (Julien-Gau et al., 2014, PMID: 24012871). As a further means of measuring possible changes in metabolism in fstr mutants and our original HGS strains, we have introduced a gut-expressed ATP FRET sensor (Mendelsohn et al., 2018, PMID: 30148842) into these backgrounds. Our preliminary data suggest that both
clk-1 and
fstr-1 fstr-2 double mutants have at most a mild impairment in ATP production in the intestine, at least in the first few days of adulthood. These results are consistent with biochemical studies on
clk-1 mutants (Braeckman et al., 1999, PMID: 10330373). Overall, our results support a role for FSTR-2 in modulation of metabolism under some conditions. Experiments to probe differences between FSTR-1 and FSTR-2 in wild-type and HGS strains are ongoing.