ADH5 Knockout HCT116 Cell Line

The choice depends on whether you are studying ADH5 (alcohol dehydrogenase 5, ADH3, GSNOR, glutathione-dependent formaldehyde dehydrogenase)'s role as the principal endogenous formaldehyde clearance enzyme or modeling AMeD syndrome and emerging research on endogenous aldehydes. The Knockout line is the standard tool for asking whether ADH5 is required for these processes — ADH5 is an atypical alcohol dehydrogenase that, unlike class I ADHs (ethanol-oxidizing ADH1/2/4), oxidizes formaldehyde-glutathione adducts (S-hydroxymethylglutathione → S-formylglutathione) and reduces S-nitrosoglutathione (GSNO); ADH5 has dual roles in endogenous formaldehyde clearance (formaldehyde is generated as a byproduct of multiple cellular processes including one-carbon metabolism and demethylation) and S-nitrosothiol homeostasis (GSNOR/GSNO reductase). Overexpression is useful for studying ADH5 gain-of-function effects. For endogenous aldehyde research, the EDITGENE ADH5 Knockout in HCT 116 is uniquely valuable — landmark studies (Dingler et al. Molecular Cell 2020; Mu et al. Nature 2021) identified ⭐⭐ ADH5/ALDH2 dual deficiency syndrome (AMeD, aplastic anemia/mental retardation/dwarfism) caused by biallelic ADH5 loss-of-function combined with ALDH2*2 East Asian variant; this established endogenous formaldehyde as a major DNA damage source. Rescue with wild-type ADH5 is the standard specificity control. The knockout is valuable for studying endogenous formaldehyde biology, formaldehyde-induced DNA damage, ICL repair coupling, GSNO/S-nitrosothiol signaling, and emerging GSNOR inhibitor research (S-nitroso-N-acetyl-DL-penicillamine, cavosonstat/N91115 in asthma trials).

Primary applications: • Endogenous formaldehyde clearance: cellular formaldehyde levels by fluorescent probes (FAP-1, others) in ADH5-null cells. • AMeD syndrome modeling: rescue with patient-derived ADH5 mutations for ⭐⭐ AMeD (aplastic anemia/mental retardation/dwarfism, ADH5/ALDH2 dual deficiency) disease modeling. • Formaldehyde-induced DNA damage: γH2AX foci kinetics analysis given the role of formaldehyde as an endogenous DNA crosslinker. • GSNO/S-nitrosothiol homeostasis: GSNO levels and S-nitrosylation of target proteins analysis given GSNOR/ADH5 activity. • GSNOR inhibitor specificity: critical genetic control for cavosonstat (N91115, GSNOR inhibitor in asthma/cystic fibrosis trials). EDITGENE recommends this model for researchers investigating endogenous formaldehyde biology, AMeD syndrome, and GSNOR-targeted therapeutic development.

Yes. ADH5 rescue experiments are well-established for endogenous aldehyde research: • Construct design: use a codon-modified ADH5 sequence with a small C-terminal tag (FLAG, HA). ADH5 has the canonical alcohol dehydrogenase architecture with active site zinc and NAD⁺/NADP⁺ binding — preserve all elements. • Catalytically-dead rescue: active site zinc-binding mutations abolish formaldehyde dehydrogenase activity. • AMeD syndrome mutation rescue: rescue with patient-derived ADH5 mutations (combined with ALDH2*2 background where appropriate) for disease modeling. • Functional readout: rescue should restore formaldehyde clearance and S-nitrosoglutathione reductase activity. HCT 116 transduces efficiently with lentivirus and supports stable rescue line generation.