Functional characterization of serotonin N-acetyltransferases from Caenorhabditis elegans and enhanced senescence tolerance in transgenic rice overexpressing CeSNAT1 via melatonin increase

• The nematode Caenorhabditis elegans has long been known to produce melatonin, but the key gene encoding serotonin N-acetyltransferase ( SNAT ) had not yet been cloned. • Neither animal-specific SNAT genes nor plant SNAT -specific homologs were identified in the C. elegans genome; however, two archaeal SNAT homologs were found. • Among the two SNAT candidates, CeSNAT1 exhibited SNAT enzyme activity in vitro . Transgenic rice overexpressing CeSNAT1 showed enhanced melatonin synthesis, which was functionally linked to senescence tolerance. • CeSNAT1 homologs are widely distributed in phylum Nematoda, including the parasitic nematode Anisakis simplex . Purified recombinant AsSNAT also displayed SNAT activity. The nematode Caenorhabditis elegans is a model invertebrate for investigating melatonin function and diverse behaviors. It synthesizes melatonin in a circadian manner, peaking at night; however, the key gene encoding serotonin N-acetyltransferase ( SNAT ) has not been cloned to date. Two archaeal SNAT homologs were identified in the C. elegans genome: CeSNAT1 , encoding a 167-amino acid (aa), and CeSNAT2 , encoding a 245-aa. Purified recombinant CeSNAT1 exhibited SNAT enzymatic activity toward serotonin ( K m = 782 μM) and 5-methoxytryptamine ( K m = 1413 μM), whereas CeSNAT2 showed no detectable SNAT activity. Heterologous overexpression of CeSNAT1 in rice enhanced melatonin synthesis and delayed senescence in transgenic plants, a phenotype commonly observed in melatonin-rich species. CeSNAT1 homologs were also widely distributed across phylum Nematoda, including the parasitic nematode Anisakis simplex . The A. simplex SNAT (AsSNAT) shared 32% aa identity with CeSNAT1. Purified recombinant AsSNAT displayed markedly lower K m values for serotonin (408 μM) and 5-methoxytryptamine (188 μM) compared with CeSNAT1, indicating a substantially higher melatonin biosynthetic capacity in the metabolically reduced parasitic nematode A. simplex than in free-living C. elegans . The de novo identification of SNAT genes in C. elegans and A. simplex paves the way for elucidating the biological roles of melatonin in nematodes from the perspective of SNAT gene regulation.