Endogenous formaldehyde scavenges cellular glutathione resulting in cytotoxic redox disruption

Abstract

25Formaldehyde (FA) is a ubiquitous endogenous and environmental metabolite that is thought to exert 26 cytotoxicity through DNA and DNA-protein crosslinking. We show here that FA can cause cellular damage 27 beyond genotoxicity by triggering oxidative stress, which is prevented by the enzyme alcohol dehydrogenase 28 5 (ADH5/GSNOR). Mechanistically, we determine that endogenous FA reacts with the redox-active thiol group 29 of glutathione (GSH) forming S-hydroxymethyl-GSH, which is metabolized by ADH5 yielding reduced GSH thus 30 preventing redox disruption. We identify the ADH5-ortholog gene in Caenorhabditis elegans and show that 31 oxidative stress also underlies FA toxicity in nematodes. Moreover, we show that endogenous GSH can protect 32 cells lacking the Fanconi Anemia DNA repair pathway from FA, which might have broad implications for 33 Fanconi Anemia patients and for healthy BRCA2-mutation carriers. We thus establish a highly conserved 34 mechanism through which endogenous FA disrupts the GSH-regulated cellular redox homeostasis that is 35 critical during development and aging. 36 37 hydroxymethylglutathione, Cancer, Genotoxicity. 38 39 BMF, liver and kidney dysfunction and early cancer onset 9 , indicating that endogenous FA can drive cancer 50 initiation and Fanconi Anemia phenotypes. 51Genotoxicity has been widely indicated as the main consequence of FA reactivity in cells 4 . However, 52 the strong reactivity of the FA carbonyl group might also affect other molecules than DNA. In vitro, the 53 spontaneous electrophilic attack of the FA carbonyl group to the thiol-group of GSH leads to the formation of 54 the covalent product S-hydroxymethyl-GSH (HSMGSH) 12 . This reaction might be strongly favored inside cells, 55where GSH levels are in the millimolar range 13 . Accordingly, ADH5 metabolizes HSMGSH yielding formate, 56 which is directed to the one-carbon cycle for nucleotide synthesis 3 (Fig. 1A). 57Considering the electrophilicity of FA and the abundance of GSH we hypothesize that the reaction 58 between FA and GSH might affect the GSH pool having detrimental biological consequences. Indeed, 59 alterations in GSH homeostasis have been reported in multiple pathologies such as hemolytic anemia, 60 diabetes, liver diseases, cystic fibrosis, neurodegeneration and cancer [14][15][16][17] . GSH not only neutralizes reactive 61 oxygen species (ROS), but can also promote chemoresistance by forming GSH-xenobiotic conjugates that are 62 pumped out of the cell via multiple resistance-associated protein transporters (MRP) 18 . To replenish 63 intracellular GSH, cells synthesize GSH in a two-step metabolic pathway centered on the rate-limiting enzyme 64 glutamate cysteine ligase (GCL), which is composed of a catalytic (GCLC) and a regulatory (GCLM) subunit, and 65 the GSH synthetase (GS) ( Fig. 1a) 18 . Cells might thus also need to maintain the balance between GSH and the 66 oxidized GSH disulfide form (GSSG) -GSH:GSSG-to limit free FA and to prevent redox disruption. 67We report here that FA toxicity is inf...

Illuminating cellular formaldehyde

“…For example, in their paper, Zhu et al showed that FAsor lights up upon blocking GSH metabolism, indicating that GSH limits intracellular FA concentrations. This result supports recent findings showing that GSH synthesis prevents FA cytotoxicity 7 . Furthermore, FAsor could be applied to screen for drugs or compounds able to modulate the cellular levels of FA with therapeutic purpose.…”

Genetically encoded formaldehyde sensors inspired by a protein intra-helical crosslinking reaction

“…Combined treatment of L-BSO and N6022 resulted in little increase with no statistical difference ( Supplementary Fig. 6b, c), indicating that ADH5 and GSH synthesis might be epistatic for the control of intracellular FA 42 .…”

“…This result confirmed that the deficiency of FA metabolism in ALDH2/ADH5 inhibited cells disrupted FA homeostasis and caused cellular FA accumulation, which may lead to cell damaging effects in related diseases. In addition, a recent report showed that inhibition of GSH synthesis with L-BSO (L-Buthionine-Sulfoximine, an inhibitor of γ-glutamylcysteine synthetase) promoted the toxicity of FA to cells, especially in those with ADH5 knockout 42 . We thus treated HeLa cells with L-BSO and N6022 alone or together, and vehicle control, respectively.…”

Effect of Microcystin-LR, Nodularin, Anatoxin-a, β-N-Methylamino-L-Alanine and Domoic Acid on Antioxidant Properties of Glutathione

“…However, knowledge of the detailed mechanisms of cyanotoxins' impact, including disrupted metabolic pathways in the context of antioxidant systems, is still scarce. It was demonstrated that some other toxins such as arsenic species or formaldehyde might interfere with a crucial antioxidant system component, glutathione (GSH), in a potentially direct reaction, which can lead to harmful effects at the cellular level [25,26]. In our recent paper, we examined the influence of CYN or its decomposition products on GSH activity in a direct reaction.…”