SIN3 maintains adaptive capacity to different diets

SIN-3 coregulator maintains adaptive capacity to different diets in Caehnorhabditis elegans through vitamin B12 | bioRxiv /* */ /* */ <!-- <!-- /*! * yepnope1.5.4 * (c) WTFPL, GPLv2 */ (function(a,b,c){function d(a){return"[object Function]"==o.call(a)}function e(a){return"string"==typeof a}function f(){}function g(a){return!a||"loaded"==a||"complete"==a||"uninitialized"==a}function h(){var a=p.shift();q=1,a?a.t?m(function(){("c"==a.t?B.injectCss:B.injectJs)(a.s,0,a.a,a.x,a.e,1)},0):(a(),h()):q=0}function i(a,c,d,e,f,i,j){function k(b){if(!o&&g(l.readyState)&&(u.r=o=1,!q&&h(),l.onload=l.onreadystatechange=null,b)){"img"!=a&&m(function(){t.removeChild(l)},50);for(var d in y[c])y[c].hasOwnProperty(d)&&y[c][d].onload()}}var j=j||B.errorTimeout,l=b.createElement(a),o=0,r=0,u={t:d,s:c,e:f,a:i,x:j};1===y[c]&&(r=1,y[c]=[]),"object"==a?l.data=c:(l.src=c,l.type=a),l.width=l.height="0",l.onerror=l.onload=l.onreadystatechange=function(){k.call(this,r)},p.splice(e,0,u),"img"!=a&&(r||2===y[c]?(t.insertBefore(l,s?null:n),m(k,j)):y[c].push(l))}function j(a,b,c,d,f){return q=0,b=b||"j",e(a)?i("c"==b?v:u,a,b,this.i++,c,d,f):(p.splice(this.i++,0,a),1==p.length&&h()),this}function k(){var a=B;return a.loader={load:j,i:0},a}var l=b.documentElement,m=a.setTimeout,n=b.getElementsByTagName("script")[0],o={}.toString,p=[],q=0,r="MozAppearance"in l.style,s=r&&!!b.createRange().compareNode,t=s?l:n.parentNode,l=a.opera&&"[object Opera]"==o.call(a.opera),l=!!b.attachEvent&&!l,u=r?"object":l?"script":"img",v=l?"script":u,w=Array.isArray||function(a){return"[object Array]"==o.call(a)},x=[],y={},z={timeout:function(a,b){return b.length&&(a.timeout=b[0]),a}},A,B;B=function(a){function b(a){var a=a.split("!"),b=x.length,c=a.pop(),d=a.length,c={url:c,origUrl:c,prefixes:a},e,f,g;for(f=0;f<d;f++)g=a[f].split("="),(e=z[g.shift()])&&(c=e(c,g));for(f=0;f<b;f++)c=x[f](c);return c}function g(a,e,f,g,h){var i=b(a),j=i.autoCallback;i.url.split(".").pop().split("?").shift(),i.bypass||(e&&(e=d(e)?e:e[a]||e[g]||e[a.split("/").pop().split("?")[0]]),i.instead?i.instead(a,e,f,g,h):(y[i.url]?i.noexec=!0:y[i.url]=1,f.load(i.url,i.forceCSS||!i.forceJS&&"css"==i.url.split(".").pop().split("?").shift()?"c":c,i.noexec,i.attrs,i.timeout),(d(e)||d(j))&&f.load(function(){k(),e&&e(i.origUrl,h,g),j&&j(i.origUrl,h,g),y[i.url]=2})))}function h(a,b){function c(a,c){if(a){if(e(a))c||(j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}),g(a,j,b,0,h);else if(Object(a)===a)for(n in m=function(){var b=0,c;for(c in a)a.hasOwnProperty(c)&&b++;return b}(),a)a.hasOwnProperty(n)&&(!c&&!--m&&(d(j)?j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}:j[n]=function(a){return function(){var b=[].slice.call(arguments);a&&a.apply(this,b),l()}}(k[n])),g(a[n],j,b,n,h))}else!c&&l()}var h=!!a.test,i=a.load||a.both,j=a.callback||f,k=j,l=a.complete||f,m,n;c(h?a.yep:a.nope,!!i),i&&c(i)}var i,j,l=this.yepnope.loader;if(e(a))g(a,0,l,0);else if(w(a))for(i=0;i (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0];var j=d.createElement(s);var dl=l!='dataLayer'?'&l='+l:'';j.src='//www.googletagmanager.com/gtm.js?id='+i+dl;j.type='text/javascript';j.async=true;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-M677548'); Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search New Results SIN-3 coregulator maintains adaptive capacity to different diets in Caehnorhabditis elegans through vitamin B12 View ORCID Profile Francesca Palladino , Cécile Bedet doi: https://doi.org/10.1101/2025.08.29.673101 Francesca Palladino 1 Laboratory of Biology and Modeling of the Cell, UMR5239 CNRS/Ecole Normale Supérieure de Lyon, INSERM U1210, UMS 3444 Biosciences Lyon Gerland, Université de Lyon , Lyon, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Francesca Palladino Cécile Bedet 1 Laboratory of Biology and Modeling of the Cell, UMR5239 CNRS/Ecole Normale Supérieure de Lyon, INSERM U1210, UMS 3444 Biosciences Lyon Gerland, Université de Lyon , Lyon, France Find this author on Google Scholar Find this author on PubMed Search for this author on this site For correspondence: cecile.bedet{at}ens-lyon.fr Abstract Full Text Info/History Metrics Supplementary material Preview PDF Abstract Diet has profound effects on metabolism and health, requiring animals to adapt their physiology to varying food sources. Here we show that the conserved SIN3 transcriptional coregulator is required in Caehnorhabditis elegans for animals to adapt to different bacterial diets. Shifting sin-3 mutant animals from the commonly used E. coli OP50 strain to the E. coli HT115 strain used for RNAi knockdown experiments has a dramatic impact on survival, leading to the lethality of mutant animals at the young adult stage. A major difference between OP50 and HT115 is vitamin B12 content, which is lower in OP50 than HT115. Feeding sin-3 mutant animals on OP50 bacteria supplemented with vitamin B12 reproduced the lethality observed on HT115. Vitamin B12 functions in two metabolic pathways: the canonical propionate breakdown pathway and the methionine/S-adenosylmethionine (met/SAM) cycle. Loss of metr-1 , encoding the methionine synthase enzyme that requires vitamin B12 as a cofactor in the met/SAM cycle repressed lethality on HT115. Conversely, complementation with metabolites of the met/SAM cycle enhanced lethality on OP50. These results show that SIN-3 is required for animals to adapt to dietary changes through B12-dependent metabolic pathways. Introduction Complex diseases arise through the interplay between genetic and environmental factors including diet ( Curran et al. 2009 ). While interactions between the host, its diet and its microbiota are essential for the health of the organism, demonstrating reproducible gene–diet interactions can be challenging. In C. elegans , numerous diet-dependent effects on animal physiology and behaviour have been reported ( Shtonda and Avery 2006 ; Soukas et al. 2009 ; Maier et al. 2010 ; Gracida and Eckmann 2013a; MacNeil et al. 2013 ; Macneil and Walhout 2013 ; Pang and Curran 2014 ; Frézal et al. 2023 ), but only a few dietgene pairs have been identified where the consequence of mutating a specific gene is only observed on specific diets ( Pang and Curran 2014 ; Verma et al. 2018 ; Amin et al. 2020 ; Nair et al. 2022 ). Here we report a diet-dependent phenotype associated with loss of the conserved SIN-3 transcriptional coregulator in C. elegans . In previous work we showed that SIN-3 is required for fertility, and plays a role in mitochondrial dynamics and metabolic flux ( Robert et al 2023 ; Giovannetti et al. 2024 ). How diet, or the observed metabolic changes relate to sin-3 mutant phenotypes and physiology was not investigated. Here we report that sin-3 mutant animals that are viable on OP50 bacteria routinely used to grow C. elegans ( Brenner 1974 ), die as young adults when fed on the HT115 bacteria utilized to elicit an RNA interference (RNAi) response. Lethality on HT115 is due to an “exploded” phenotype resulting from extrusion of the intestine through the vulva. One well described difference between these two bacterial strains is vitamin B12 content, which is lower in OP50 than HT115 ( Watson et al. 2013 ; Watson et al. 2014 ; Watson et al. 2016 ; Revtovich et al. 2019 ; Nair et al. 2022 ), and we show that complementing OP50 with vitamin B12 reproduces the “exploded” phenotype and lethality of sin-3 mutants on HT115. As in humans, in C. elegans vitamin B12 is required in two metabolic pathways, the canonical mitochondrial propionate breakdown pathway, and one-carbon (1C) metabolism composed of the methionine/S-adenosylmethionine (met/SAM) and folate cycles. In the canonical propionate breakdown pathway, vitamin B12 acts as a co-factor for methyl malonyl-CoA mutase (MUT, MMCM-1 in C. elegans ) that catalyzes the conversion of methylmalonyl-CoA to succinyl-CoA. In the methionine cycle, it is a cofactor for methionine synthase (MS, METR-1 in C. elegans ) that synthesizes methionine from homocysteine (Watson et al. 2013; Watson et al. 2014; Watson et al. 2016; Green et al. 2017). We show that RNAi knock-down of metr-1 , but not mmcm-1 , suppresses the lethality of sin-3 mutant animals on the B12-replete HT115 diet, indicating that the activity of the met/SAM cycle contributes to lethality. Supplementation with 1C cycle metabolites further showed that the met/SAM, but not the folate cycle, contributes to lethality in sin-3 mutant animals on the vitamin B12-depleted OP50 strain. Although the vitamin B12-dependent activity of the canonical propionate breakdown pathway does not contribute to lethality in sin-3 mutants, these animals are more sensitive to propionate on the OP50 diet, possibly due to reduced expression in these animals of genes in the alternative propionate shunt pathway active under low vitamin B12 conditions (Watson et al. 2016). Altogether these results suggest that metabolic rewiring in the absence of sin-3 renders animals sensitive to vitamin B12, identifying a role for SIN-3 in adaptation to dietary changes. Results Dietary vitamin B12 affects sin-3 viability The first hint that food source can impact the physiology of sin-3 mutant animals came from the observation that a high proportion of animals carrying the sin-3(tm1276) hypomorphic mutation ( Robert et al. 2023 ) and fed on HT115, but not OP50 bacteria, died as young adults with an exploded vulva resulting in extrusion of the intestine ( Figure 1A , right panel). The exploded phenotype, which is one of multiple pathologies observed in aged wild-type animals, has also been referred to as “age-associated vulval integrity defects” phenotype (AVID) (Leiser et al. 2016). Here we refer to the phenotype we observe in young animals as “exploded”, without any reference to the underlying cause, which was not investigated in this study. To quantify the exploded phenotype, synchronized animals at the L1 stage were grown on plates seeded with either OP50 or HT115 bacteria, and visually scored when they reached day 3 of adulthood. No diet-dependent difference was observed in wildtype N2 adult animals, which as expected never showed the exploded phenotype at this stage (Leiser et al. 2016, Figure 1B ). sin-3 mutant animals grown on OP50 developed normally to adulthood, with fewer than 5% dying from a ruptured vulva. In contrast, when grown on HT115 bacteria, 28% of day-3 mutant adults died with this phenotype. The number of exploded sin-3 mutant animals on HT115 peaked at day 5 of adulthood, at which time point we sometimes observed more than 75% lethality, while wildtype animals were 100% viable at this stage (Figure S1A). We chose the day 3 time point for all subsequent experiments in order to be able to measure any potential enhancement of this phenotype. Download figure Open in new tab Figure 1. Vitamin B12 levels are responsible for the increased lethality of sin-3 mutant animals on HT115. ( A) Representative images of worms at day 3 of adulthood for wildtype (left panel) and sin-3 mutant animals (middle and right panels) fed on HT115 bacteria. The right panel shows a sin-3 mutant adult with an exploded vulva phenotype resulting in extrusion of the intestine. (B) Lethality due to exploded vulva of sin-3 mutant animals is significantly increased when worms are grown on HT115 bacteria, or on OP50 supplemented with vitamin B12 (25 nM). n = total number of animals scored. Circles represent independent plates with 15-64 animals. A two-proportions two-tailed z-test was carried out to perform pairwise comparison. *** P<0.05. OP50 and HT115 bacteria differ in both the amount and composition of nutrients and metabolites ( Brooks et al. 2009 ; Reinke et al. 2010 ). Because of this, these two diets differentially affect nearly every aspect of worm biology and physiology, including development, metabolism, behavior, and aging ( Jeong et al. 2009 ; Gracida and Eckmann 2013 ; Reinke et al. 2010 ; Yen and Curran 2016 ; Stuhr and Curran 2020 ; Frézal et al. 2023 ). One of the major differences identified between these two strains is the level of vitamin B12. C. elegans acquires vitamin B12 from its bacterial food source. However, the E. coli strain OP50 poorly expresses TonB ( Bassford et al. 1976 ), a transporter required for vitamin B12 uptake from the environment, causing it to be relatively deficient in vitamin B12 compared to HT115 ( Watson et al. 2013 ; Watson et al. 2014 ; Revtovich et al. 2019 ; Nair et al. 2022 ). Supplementation of OP50-seeded plates with 25 nM cobalamin (vitamin B12) significantly increased the proportion of sin-3 exploded animals from less than 5% to 20% ( Figure 1B ). Lethality of sin-3 mutant animals was observed at concentrations as low as 3 nM (Figure S1B). Similarly low concentrations of B12 have been show to impact C. elegans physiology ( McDonagh et al. 2022 ; Qin et al. 2022 ). These results are consistent with vitamin B12 levels being responsible for the increased lethality observed on HT115. Activity of the vitamin B12-dependent enzyme metr-1 contributes to the exploded phenotype of sin-3 mutant animals Vitamin B12 is an important co-factor in two metabolic pathways: the mitochondrial propionate breakdown pathway that prevents the toxic accumulation of propionate, and 1C metabolism formed by the met/SAM and folate cycles ( Figure 2A ). In the canonical propionate breakdown pathway, vitamin B12 acts as a co-factor for MMCM-1, leading to the production of succinyl-CoA. In the met/SAM cycle it serves as a co-factor for METR-1 ( Watson et al. 2014 ; Watson et al. 2016 ; Green et al. 2017 ). We found that RNAi inactivation of metr-1 rescued sin-3 mutant lethality on HT115, with the percent of exploded animals significantly decreasing to less than 5% ( Figure 2B ). mmcm-1 knockdown by contrast had no significant impact on the sin-3 exploded phenotype. RT-qPCR confirmed a similar efficacy of RNAi knock-down in both wildtype and mutant backgrounds for each independent replica (Figure S2 A and B). These results indicate that vitamin B12 acts through metr-1 in the met/SAM cycle to give rise to the exploded phenotype in sin-3 mutant animals. Download figure Open in new tab Figure 2. Methionine cycle perturbation is involved in survival of sin-3 mutant animals. (A) Cartoon showing C. elegans one-carbon and propionate breakdown metabolic pathways, and link of Met/SAM cycle to PC synthesis. The folate cycle was simplified to illustrate the role of vitamin B12 in tetrahydrofolate (THF) synthesis. Enzymes are written in red. Red stars correspond to metabolite tested in figure C. (B) Inactivation of metr-1 , but not mmcm-1 by RNAi rescued lethality of sin-3 mutant animals on HT115. Four independent experiments were performed. A two-proportions two-tailed z-test was carried out to perform pairwise comparison. *** P<0.05. (C) Supplementation with methionine (4 mM), SAM (500 µM or 100µM) or choline (40 mM), but not folinic acid (100 µM) increases lethality of sin-3 mutant animals. n = total number of animals scored. A two-proportions two-tailed z-test was carried out to perform pairwise comparison, *** P<0.05. 1C metabolites affect lethality in sin-3 mutants Having shown that vitamin B12 acts through metr-1 in the met/SAM cycle to negatively impact survival of sin-3 mutant animals, we carried out additional experiments to identify relevant metabolites in this pathway. In 1C metabolism, 5-methyltetrahydrofolate (5-methyl-THF) acts as a methyl donor for the synthesis of methionine from homocysteine through the activity of METR-1/MS. Methionine is then converted into SAM, the main methyl donor in cells required for phosphatidylcholine (PC) synthesis ( Ducker and Rabinowitz 2017 ; Ye et al. 2017 and Figure 2A ), as well as DNA, RNA, histone and protein methylation ( Chiang et al. 1996 ). We asked whether feeding sin-3 mutant animals on OP50 bacteria supplemented with different 1C metabolites reproduced the exploded phenotype and lethality observed on HT115. Specifically, we tested folinic acid, a derivative of tetrahydrofolic acid that is converted into the different metabolites to enhance the folate cycle, as well as methionine and SAM to enhance the met/SAM cycle ( Figure 2A ). Both methionine and SAM enhanced the exploded phenotype of mutant animals on OP50 ( Figure 2C ). The methylation-dependent pathway represents only one of the 2 avenues to synthesize PC ( DeLong et al. 1999 ), which can also be synthesized from choline ( Figure 2A ). Choline supplementation also increased lethality in sin-3 mutants ( Figure 2C ), suggesting that increased PC synthesis by either pathway may be deleterious in the context of SIN-3 depletion. Altogether, these results suggest that in the absence of SIN-3, METR-1 activity contributes to the exploded phenotype through methionine, SAM and PC. By contrast, folinic acid supplementation had no effect on the survival of sin-3 mutants, suggesting that this branch of 1C metabolism is not implicated ( Figure 2C ). Increased propionate sensitivity in sin-3 mutant animals RNAi knockdown of mmcm-1 , which requires vitamin B12 as a cofactor in the canonical mitochondrial propionate breakdown pathway ( Figure 2A ), had no effect on HT115 induced lethality, suggesting that this pathway is not implicated. However, in C. elegans an alternate propionate breakdown pathway, or shunt, is used under low vitamin B12 dietary conditions, or when flux through the canonical pathway is genetically perturbed ( Watson et al. 2013 ; Watson et al. 2014 ; Watson et al. 2016 and Figure 2A ). We were intrigued by the observation that three of the four conserved downstream genes in this pathway were downregulated in our previous transcriptomic analysis on sin-3 mutants ( Robert et al. 2023 ): hach-1 , a 3-hydroxyisobutyryl-CoA hydrolase, hphd-1 , a hydroxyacid-oxoacid transhydrogenase, and alh-8 , a decarboxylating dehydrogenase ( Watson et al. 2016 ) (Figure S5A). Based on this observation, we tested whether the sensitivity of sin-3 mutant animals to propionate is altered. Survival assays revealed that sin-3 mutants are significantly more sensitive to propionate than wild type, with an LD50 of 27.9 mM compared to 45.2 mM for wild type ( Figure 3A and B ). The first gene of the propionate shunt, the acyl-CoA dehydrogenase acdh-1 , is differentially expressed depending on the vitamin B12/propionate axis: its transcript levels are very low when vitamin B12 is high (as on HT115) and the canonical breakdown pathway is active, and increase several hundred-fold when propionate accumulates under low vitamin B12 levels (as on OP50) ( Watson et al. 2013 ; Watson et al. 2014 ). We found that acdh-1 induction occurs normally on OP50 in sin-3 mutants, indicating proper activation of the shunt pathway (Figure S4). Together, these results suggest that although sin-3 mutant animals can initiate this pathway, downstream components — hach-1, hphd-1 , and alh-8 — may not be fully activated, rendering the pathway less efficient. Interestingly, our previously published ChIP-seq data ( Beurton et al. 2019 ) revealed the presence of SIN-3 at the promoter region of these same genes (Figure S5B), suggesting a possible direct role in their regulation. Because the propionate shunt pathway may help animals adapt to changes in diet and vitamin B12 content in the wild, these results suggest an additional way in which loss of sin-3 may alter the function of metabolic networks in response to diet. Download figure Open in new tab Figure 3. sin-3 mutant animals are more sensitive to propionate than wildtype (A) Dose-response curves showing percent survival of sin-3 and wildtype on increasing propionate concentrations. Three independent experiments were performed and the resulting curves were fitted using HelpersMG package in R. (B) Average LD50 and standard deviation of data shown in (A). Unpaired student’s T tests were used to calculate p-values, ** P<0.01. Loss of sin-3 reshapes the mitochondrial network independently of the canonical DRP-1 pathway Among its many functions in animal physiology, vitamin B12 has been implicated in the regulation of mitochondrial dynamics through multiple metabolic mechanisms that can either stabilize or disrupt mitochondrial networks ( Revtovich et al. 2019 ; Wei and Ruvkun 2020 ). Specifically, recent work demonstrated that mitochondrial dynamics can be restored in mutant animals defective in the canonical DRP-1 mediated fission pathway ( Campbell and Zuryn 2024 ) by growth on B12-depleted OP50, or by reduced activity of the B12-dependent enzyme METR-1, revealing a metabolism-driven and DRP-1– independent pathway for mitochondrial fission ( Wei and Ruvkun 2020 ). Because in this context B12 depletion suppressed defects in drp-1 mutants that are defective in fission, it is unlikely to improve mitochondria morphology in sin-3 mutants, which already have increased fission ( Giovannetti et al. 2024 ). However, because of the common link to B12-dependent metabolism and met/SAM cycle activity, we asked whether the increased fragmentation in sin-3 mutants may also be mediated by a DRP-1-independent pathway by looking at the effect of drp-1 inactivation on mitochondria dynamics in these mutants. Because sin-3; drp-1(tm1108) double mutants are sterile, we used RNAi-mediated knockdown of drp-1 . To enable mitochondrial scoring while avoiding the exploded phenotype observed in sin-3 mutants on HT115, we performed RNAi using the iOP50 strain ( Neve et al. 2020 ). As expected, drp-1 (RNAi) reduced fertility in wild-type animals (Kanazawa 2008; Machiela et al. 2021 ), while sin-3; drp-1 (RNAi) animals were fully sterile, consistent with the genetic double mutant and confirming effective drp-1 knockdown (Figure S3A, B). Mitochondrial morphology was therefore examined in day 1 sterile animals. Consistent with prior reports, microscopy analysis of mitochondria morphology in muscle cells using the TOMM-20::RFP reporter revealed aggregated mitochondrial networks connected by thin tubules in drp-1 depleted cells (Dong et al. 2018; Figure S3C) ( Dong et al. 2018 ; Figure S3C). In contrast, sin-3 mutants displayed increased mitochondrial fragmentation or aberrant interconnections, as previously described ( Giovannetti et al. 2024 ). Notably, mitochondrial morphology in sin-3; drp-1 (RNAi) animals showed no evidence of rescue: mitochondria remained highly fragmented and, in some animals, appeared even more disorganized or excessively interconnected than in either single perturbation alone (Figure S3C). Although the severity and variability of mitochondrial abnormalities prevented reproducible quantitative analysis, these observations were consistent across independent experiments. Together, these results suggest that the mitochondrial fragmentation observed upon loss of sin-3 does not require the canonical DRP-1–dependent fission pathway. Whether DRP-1-independent changes in mitochondrial dynamics in sin-3 mutants are linked to metabolism and the observed diet-dependent lethality remains to be established. Discussion In response to changing environments, metabolic networks can be rewired through the use of alternate enzymes to adjust pathway flux, thereby maintaining organismal homeostasis. Our results suggest that in C. elegans , the SIN-3 coregulator may play a role in maintaining adaptive capacity on different sources of food. We found that sin-3 mutant animals grown on vitamin B12-replete HT115 bacteria die with an exploded vulva, a phenotype that we only rarely observed on OP50. Knock-down of the vitamin B12-dependent METR-1 methionine-synthase in mutant animals suppressed the exploded phenotype and adult lethality on HT115. Conversely, methionine or SAM supplementation enhanced lethality on the vitamin B12-depleted OP50 diet. Together, these results suggest that met/SAM cycle activity contributes to lethality in the absence of SIN-3. Met/SAM cycle activity is tightly regulated by a gene regulatory network to support organismal homeostasis (Giese et al. 2020), and either impaired or excessive activity is deleterious, reflecting the role of SAM as the major methyl donor in cells (Finkelstein 1990). Our results indicate that sin-3 mutants are particularly sensitive to vitamin B12-dependent 1C metabolism, specifically the channeling of methionine and SAM into the met/SAM cycle. A specific vitamin B12 diet-dependent vulnerability was previously identified in a C. elegans model of a neomorphic mutations in isocitrate dehydrogenase ( idh-1neo ). In humans idh1neo causes increased levels of cellular D-2-hydroxyglutarate (D-2HG), a proposed oncometabolite (Losman and Kaelin 2013). Increased vitamin B12 levels enhanced embryonic lethality of C. elegans idh-1neo mutants through increased utilization of the met/SAM cycle, resulting in a decrease in the cellular 1C pool. In this context, supplementation with alternate sources of 1C donors suppressed lethality ( Ponomarova et al., 2024 ). In sin- 3 mutants by contrast, supplementation with the 1C donors methionine or SAM enhanced the exploded phenotype and lethality on OP50. Several explanations could account for this effect. In sin-3 mutants, both chromatin landscape and transcription patterns are significantly altered, with evidence of changes in histone modification states and widespread misregulation of gene expression ( Robert et al. 2023 ). Elevated SAM levels may intensify these disruptions, for example by inducing further changes in histone methylation, which in turn could profoundly influence transcriptional activity (Mentch et al. 2015). Alternatively, or in addition, increasing the cellular 1C pool may increase flux to the polyamine synthesis pathway that also uses SAM as a substrate. Like SAM synthesis, polyamine biosynthesis is tightly controlled, with changes in polyamine metabolism linked to human diseases including cancer and diabetes (Nakanishi and Cleveland 2021). Polyamine biosynthesis is significantly increased in sin-3 mutant animals ( Giovannetti et al, 2024 ), and a further increase may potentially have deleterious consequences on animal health. In this study we focused on vitamin B12 dependent metabolic pathways whose function is altered in sin-3 mutants, notably the met/SAM cycle. However, both loss of sin-3 and vitamin B12 impact mitochondria homeostasis, and may impact metabolism and animal physiology in additional ways (Janssen et al. 2019; Revtovich et al. 2019 ; Wei and Ruvkun 2020 ; Giovannetti et al, 2024 ). While vitamin B12 depletion re-established normal mitochondria dynamics in drp-1 /DRP-1 mutants defective in fission ( Wei and Ruvkun 2020 ), its effect, if any, on sin-3 mutants in which fission is already increased is unclear. However, the finding that increased mitochondrial fission in the absence of SIN-3 occurs independently of the canonical DRP-1 pathway suggests a potential link to alternative, metabolism-dependent mechanisms of mitochondrial dynamics maintenance ( Wei & Ruvkun, 2020 ). In conclusion, our results identify SIN-3 as a key regulator that rewires the C. elegans metabolic network in response to diet and vitamin B12 levels. Because gene-diet interactions play important role in the pathophysiology of human diseases ( Singar et al. 2024 ), our study underscores the importance of studies in model organisms to unravel they underlying mechanisms, with potential implications for diet-based interventions. Materiel and methods Strains C. elegans strains were cultured on Nematode Growth Medium (NGM) agarose plates with Escherichia coli OP50 or HT115 and incubated at 20°C unless otherwise indicated. The following strains were used: wild type N2 Bristol, PFR590 sin-3(tm1276)I , EN7714 krSi134[myo-3p::tomm-20Nter::wrmScarlet] , PFR754 krSi134[myo-3p::tomm-20Nter::wrmScarlet]; sin-3(tm1276)I , VL749 wwIs24 [acdh-1p::GFP + unc-119(+)] , PFR761 wwIs24[acdh-1p::GFP + unc-119(+)]; sin-3(tm1276)I . PFR761 was obtained by crossing sin-3(tm1276) hermaphrodites with acdh-1p ::GFP males. Effect of vitamin B12 on sin-3 mutant animals on OP50 Methylcobalamine (vitamin B12 analog, #M9756, Sigma-Aldrich) was added at 25 nM to liquid NGM before pouring plates, which were then seeded with either OP50 or HT115 bacterial culture. Wildtype and sin-3(tm1276) worms were synchronized by egg laying on OP50 plates with or without vitamin B12, or on unsupplemented HT115, and grown at 20°C. Animals with exploded vulvas were scored at day 3 of adulthood. For pairwise comparison, a two-proportions two-tailed z-test was carried out in R with the prop.test() function. The corresponding script is available upon request. Metabolite assays All metabolites were added in liquid NGM before pouring the plates at the following final concentrations: L-methionine (#A1340,0100-ITW, ITW reagents) 4 mM, S-(5′-Adenosyl)-L-methionine chloride dihydrochloride (SAM) (#A7007-25MG Sigma-Aldrich) 500 µM, choline chloride (#1102900500 Thermo Scientific) 40 mM, and folinic acid calcium salt hydrate (#F7878-100MG Sigma-Aldrich) 100 µM. For scoring the exploded vulva phenotype, wildtype and sin-3(tm1276) worms were synchronized by egg laying on OP50 plates with or without supplementation and grown at 20°C until day 3 of adulthood. Supplements were therefore provided throughout development. For each metabolite, at least 4 independent experiments were performed for sin-3 mutant animals and 3 for wild-type animals. For pairwise comparison, a two-proportions two-tailed z-test was carried out in R with the prop.test() function. The corresponding script is available upon request. RNAi experiments on HT115 HT115 expressing either metr-1 or mmcm-1 RNAi or empty vector were grown in liquid LB with ampicillin (1 µg/ ml) and tetracycline (12.5 µg/ ml) overnight. 220 µl of bacterial culture was then plated on NGM agar plates with IPTG (1 mM) and plates allowed to dry for 18 to 20 hours at room temperature to induce RNAi. Three wildtype or sin-3 mutant L4 animals (P0) were transferred to HT115 control (empty vector) or metr-1 / mmcm-1 RNAi plates and allowed to develop into egg-laying adults. Three or four F1 gravid adult were transferred to new RNAi or control plates and allowed to lay eggs for 6-7 h at 20°C. F2 adults were transferred to several new RNAi plates to avoid starvation and allowed to develop to day 4 of adulthood before scoring exploded vulvas. F o u r independent RNAi experiments were performed. For each of them, RNA was extracted from F1 worms and used in RT-qPCR experiments to check RNAi efficiency as described in Giovanetti et al 2024 (see supplemental Materiel& Methods). Propionate survival assay Wildtype or sin-3 mutant animals were collected in M9 buffer from NGM plates and passed through a 40 µM cell strainer (Clearline #141308C). Gravid adults retained on the filter were recovered in 2-3 ml of M9 in a 50 ml conical tube and allowed to lay eggs overnight at 20°C with agitation (220 rpm) without food to obtain hatched animals arrest at the L1 larval stage. L1 were separated from adults by filtering through a 10 µM cell strainer (Pluriselect #43-50010-01), and approximatively 40-50 starved L1 were transferred to NGM plates containing 0, 20 mM, 40 mM and 60 mM propionic acid. Three days later, survivors were counted. Three independent experiments were performed and a dose response curve was generated for each experiment using the LD50 function from HelpersMG package ( https://search.r-project.org/CRAN/refmans/HelpersMG/html/LD50.html ). A global dose response curve combining all three experiments was generated and presented in Figure 3A . The barplot presented in Figure 3B was generated using the LD50 of each experiment and unpaired student’s T tests were used to calculate p-values. Competing Interests No competing interests declared Funding This work was supported by ANR (Agence Nationale de la Recherche) grant N° 19-CE12-0025-01 and the Centre National de la Recherche Scientifique. Data and resource availability All relevant data and details of resources can be found within the article and its supplementary information. GSE accession numbers are : GSE227498 for sin-3 transcriptomics.and GSE114715 for sin-3 ChIP-seq Acknowledgements We acknowledge the SFR Biosciences (UAR3444/CNRS, US8/Inserm, ENS de Lyon, UCBL): PLATIM. Thanks to Paul Mareshal for help in microscopy image acquisition. Funder Information Declared ANR (Agence Nationale de la Recherche) , 19-CE12-0025-01 Centre National pour la Recherche Scientifique (CNRS) Footnotes Figures and text have been revised. More replicas have been added to figures where only 3 were available in previous version. Title was modified and Materiel and Method section was clarified References ↵ Murphy CT Amin MR et al. 2020 . A novel gene-diet interaction promotes organismal lifespan and host protection during infection via the mitochondrial UPR Murphy CT , editor. PLoS Genet . 16 ( 12 ): e1009234 . doi: 10.1371/journal.pgen.1009234 OpenUrl CrossRef PubMed ↵ Bassford PJ , Bradbeer C , Kadner RJ , Schnaitman CA . 1976 . Transport of vitamin B12 in tonB mutants of Escherichia coli . J Bacteriol . 128 ( 1 ): 242 – 247 . doi: 10.1128/jb.128.1.242-247.1976 OpenUrl Abstract / FREE Full Text ↵ Beurton F et al. 2019 . Physical and functional interaction between SET1/COMPASS complex component CFP-1 and a Sin3S HDAC complex in C . elegans. Nucleic Acids Res . 47 ( 21 ): 11164 – 11180 . doi: 10.1093/nar/gkz880 OpenUrl CrossRef PubMed ↵ Brenner S. 1974 . The Genetics of Caenorhabditis elegans . Genetics . 77 ( 1 ): 71 – 94 OpenUrl Abstract / FREE Full Text ↵ Melov S Brooks KK , Liang B , Watts JL . 2009 . The Influence of Bacterial Diet on Fat Storage in C. elegans Melov S , editor. PLoS ONE . 4 ( 10 ): e7545 . doi: 10.1371/journal.pone.0007545 OpenUrl CrossRef PubMed ↵ Campbell D , Zuryn S. 2024 . The mechanisms and roles of mitochondrial dynamics in C. elegans . Seminars in Cell & Developmental Biology . 156 : 266 – 275 . doi: 10.1016/j.semcdb.2023.10.006 OpenUrl CrossRef PubMed Chen TY et al. 2024 . Mitochondrial S-adenosylmethionine deficiency induces mitochondrial unfolded protein response and extends lifespan in Caenorhabditis elegans . Aging Cell . 23 ( 4 ): e14103 . doi: 10.1111/acel.14103 OpenUrl CrossRef PubMed ↵ Chiang PK et al. 1996 . S-Adenosylmethionine and methylation . FASEB J . 10 ( 4 ): 471 – 480 OpenUrl CrossRef PubMed Web of Science ↵ Curran SP , et al. 2009 . A soma-to-germline transformation in long-lived Caenorhabditis elegans mutants . Nature . 459 ( 7250 ): 1079 – 84 OpenUrl CrossRef PubMed Web of Science ↵ DeLong CJ , Shen Y-J , Thomas MJ , Cui Z. 1999 . Molecular Distinction of Phosphatidylcholine Synthesis between the CDP-Choline Pathway and Phosphatidylethanolamine Methylation Pathway . Journal of Biological Chemistry . 274 ( 42 ): 29683 – 29688 . doi: 10.1074/jbc.274.42.29683 OpenUrl Abstract / FREE Full Text ↵ Dong L et al. 2018 . Reversible and long-term immobilization in a hydrogel-microbead matrix for high-resolution imaging of Caenorhabditis elegans and other small organisms Dupuy D, editor . PLoS ONE . 13 ( 3 ): e0193989 . doi: 10.1371/journal.pone.0193989 OpenUrl CrossRef PubMed ↵ Ducker GS , Rabinowitz JD . 2017 . One-Carbon Metabolism in Health and Disease . Cell Metabolism . 25 ( 1 ): 27 – 42 . doi: 10.1016/j.cmet.2016.08.009 OpenUrl CrossRef PubMed ↵ Frézal L et al. 2023 . Genome-wide association and environmental suppression of the mortal germline phenotype of wild C. elegans . EMBO Reports . 24 ( 12 ): e58116 . doi: 10.15252/embr.202358116 OpenUrl CrossRef PubMed ↵ Giovannetti M , et al. 2024 SIN-3 transcriptional coregulator maintains mitochondrial homeostasis and polyamine flux . iScience . 27 ( 5 ): 109789 . doi: 10.1016/j.isci.2024.109789 OpenUrl CrossRef PubMed ↵ Gracida X , Eckmann CR . 2013 . Fertility and germline stem cell maintenance under different diets requires nhr-114/HNF4 in C. elegans . Curr Biol . 23 ( 7 ): 607 – 613 . doi: 10.1016/j.cub.2013.02.034 OpenUrl CrossRef PubMed ↵ Green R et al. 2017 . Correction: Vitamin B12 deficiency . Nat Rev Dis Primers . 3 ( 1 ): 17054 . doi: 10.1038/nrdp.2017.54 OpenUrl CrossRef PubMed ↵ Jeong PY , Kwon MS , Joo HJ , Paik YK . 2009 . Molecular time-course and the metabolic basis of entry into dauer in Caenorhabditis elegans . PloS one . 4 ( 1 ): e4162 OpenUrl CrossRef PubMed Leiser SF et al. 2016 . Age-associated vulval integrity is an important marker of nematode healthspan . AGE . 38 ( 5–6 ): 419 – 431 . doi: 10.1007/s11357-016-9936-8 OpenUrl CrossRef PubMed ↵ Machiela E et al. 2021 . Targeting Mitochondrial Network Disorganization is Protective in C. elegans Models of Huntington’s Disease . Aging and disease . 12 ( 7 ): 1753 . doi: 10.14336/AD.2021.0404 OpenUrl CrossRef PubMed ↵ MacNeil LT et al. 2013 . Diet-induced developmental acceleration independent of TOR and insulin in C. elegans . Cell . 153 ( 1 ): 240 – 252 . doi: 10.1016/j.cell.2013.02.049 OpenUrl CrossRef PubMed Web of Science ↵ Macneil LT , Walhout AJ . 2013 . Food, pathogen, signal: The multifaceted nature of a bacterial diet . Worm . 2 ( 4 ): e26454 . doi: 10.4161/worm.26454 OpenUrl CrossRef PubMed ↵ Maier W , Adilov B , Regenass M , Alcedo J. 2010 . A neuromedin U receptor acts with the sensory system to modulate food type-dependent effects on C. elegans lifespan . PLoS Biol . 8 ( 5 ): e1000376 . doi: 10.1371/journal.pbio.1000376 OpenUrl CrossRef PubMed ↵ Fay D McDonagh A , Crew J , Van Der Linden AM . 2022 . Dietary vitamin B12 regulates chemosensory receptor gene expression via the MEF2 transcription factor in Caenorhabditis elegans Fay D , editor. G3 . 12 ( 6 ): jkac107 . doi: 10.1093/g3journal/jkac107 OpenUrl CrossRef ↵ Nair T et al. 2022 . Adaptive capacity to dietary Vitamin B12 levels is maintained by a gene-diet interaction that ensures optimal life span . Aging Cell . 21 ( 1 ): e13518 . doi: 10.1111/acel.13518 OpenUrl CrossRef PubMed ↵ Neve IAA et al. 2020 . Escherichia coli Metabolite Profiling Leads to the Development of an RNA Interference Strain for Caenorhabditis elegans . G3 Genes|Genomes|Genetics . 10 ( 1 ): 189 – 198 . doi: 10.1534/g3.119.400741 OpenUrl Abstract / FREE Full Text ↵ Pang S , Curran SP . 2014 . Adaptive capacity to bacterial diet modulates aging in C. elegans . Cell Metab . 19 ( 2 ): 221 – 231 . doi: 10.1016/j.cmet.2013.12.005 OpenUrl CrossRef PubMed ↵ Ponomarova O , et al. 2024 . idh-1 neomorphic mutation confers sensitivity to vitamin B12 in Caenorhabditis elegans . Life Sci Alliance . 7 ( 10 ): e202402924 . doi: 10.26508/lsa.202402924 OpenUrl Abstract / FREE Full Text ↵ Qin S et al. 2022 . Early-life vitamin B12 orchestrates lipid peroxidation to ensure reproductive success via SBP-1/SREBP1 in Caenorhabditis elegans . Cell Reports . 40 ( 12 ): 111381 . doi: 10.1016/j.celrep.2022.111381 OpenUrl CrossRef PubMed ↵ Reinke SN , Hu X , Sykes BD , Lemire BD . 2010 . Caenorhabditis elegans diet significantly affects metabolic profile, mitochondrial DNA levels, lifespan and brood size . Molecular Genetics and Metabolism . 100 ( 3 ): 274 – 282 . doi: 10.1016/j.ymgme.2010.03.013 OpenUrl CrossRef PubMed ↵ Revtovich AV , Lee R , Kirienko NV . 2019 . Interplay between mitochondria and diet mediates pathogen and stress resistance in Caenorhabditis elegans Garsin DA, editor . PLoS Genet . 15 ( 3 ): e1008011 . doi: 10.1371/journal.pgen.1008011 OpenUrl CrossRef PubMed ↵ Robert VJ , Caron M , Gely L , Adrait A , Pakulska V , Couté Y , Chevalier M , Riedel CG , Bedet C , Palladino F. 2023 . SIN-3 acts in distinct complexes to regulate the germline transcriptional program in Caenorhabditis elegans . Development . 150 ( 21 ): dev201755 . doi: 10.1242/dev.201755 .. OpenUrl CrossRef ↵ Shtonda BB , Avery L. 2006 . Dietary choice behavior in Caenorhabditis elegans . Journal of Experimental Biology . 209 ( 1 ): 89 – 102 . doi: 10.1242/jeb.01955 OpenUrl Abstract / FREE Full Text ↵ Singar S , Nagpal R , Arjmandi BH , Akhavan NS . 2024 . Personalized Nutrition: Tailoring Dietary Recommendations through Genetic Insights . Nutrients . 16 ( 16 ): 2673 . doi: 10.3390/nu16162673 OpenUrl CrossRef PubMed ↵ Soukas AA et al. 2009 . Rictor/TORC2 regulates fat metabolism, feeding, growth, and life span in Caenorhabditis elegans . Genes Dev . 23 ( 4 ): 496 – 511 . doi: 10.1101/gad.1775409 OpenUrl Abstract / FREE Full Text ↵ Stuhr NL , Curran SP . 2020 . Bacterial diets differentially alter lifespan and healthspan trajectories in C. elegans . Commun Biol . 3 ( 1 ): 653 . doi: 10.1038/s42003-020-01379-1 OpenUrl CrossRef PubMed ↵ Lee SS Verma S , Jagtap U , Goyala A , Mukhopadhyay A. 2018 . A novel gene-diet pair modulates C. elegans aging Lee SS , editor. PLoS Genet . 14 ( 8 ): e1007608 . doi: 10.1371/journal.pgen.1007608 OpenUrl CrossRef PubMed ↵ Watson E et al. 2013 . Integration of Metabolic and Gene Regulatory Networks Modulates the C. elegans Dietary Response . Cell . 153 ( 1 ): 253 – 266 . doi: 10.1016/j.cell.2013.02.050 OpenUrl CrossRef PubMed ↵ Watson E et al. 2014 . Interspecies Systems Biology Uncovers Metabolites Affecting C. elegans Gene Expression and Life History Traits . Cell . 156 ( 6 ): 1336 – 1337 . doi: 10.1016/j.cell.2014.02.036 OpenUrl CrossRef PubMed ↵ Watson E et al. 2016 . Metabolic network rewiring of propionate flux compensates vitamin B12 deficiency in C. elegans . eLife . 5 : e17670 . doi: 10.7554/eLife.17670 OpenUrl CrossRef PubMed ↵ Wei W , Ruvkun G. 2020 . Lysosomal activity regulates Caenorhabditis elegans mitochondrial dynamics through vitamin B12 metabolism . Proc Natl Acad Sci USA . 117 ( 33 ): 19970 – 19981 . doi: 10.1073/pnas.2008021117 OpenUrl Abstract / FREE Full Text ↵ Ye C et al. 2017 . A Metabolic Function for Phospholipid and Histone Methylation . Molecular Cell . 66 ( 2 ): 180 - 193.e8 . doi: 10.1016/j.molcel.2017.02.026 OpenUrl CrossRef PubMed ↵ Yen CA , Curran SP . 2016 . Gene-diet interactions and aging in C. elegans . Experimental Gerontology . 86 : 106 – 112 . doi: 10.1016/j.exger.2016.02.012 OpenUrl CrossRef PubMed View the discussion thread. Back to top Previous Next Posted February 23, 2026. 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