NSUN6 is an S-adenosyl-L-methionine-dependent methyltransferase that catalyzes 5-methylcytosine (m5C) modification at the C72 position of specific tRNAs, particularly tRNA(Thr) and tRNA(Cys) 1. The enzyme recognizes a complex network of tRNA structural elements including the 3'-CCA terminus, discriminator base U73, and specific base pairs in the acceptor and D-stem regions 1. Beyond its canonical tRNA function, NSUN6 also catalyzes m5C modification of mRNAs containing CUCCA motifs in stem-loop structures, with m5C-modified mRNAs stabilized through preferential binding by m5C readers ALYREF and YBX proteins 2. Dysregulation of NSUN6 is implicated in multiple cancers: elevated NSUN6 promotes radioresistance in cervical cancer via the NSUN6/ALYREF-m5C-NDRG1 axis 3, enhances colon adenocarcinoma progression through METTL3 m5C modification 4, and facilitates breast cancer cell migration 2. Pan-cancer analysis reveals NSUN6 overexpression in eight cancer types correlates with altered prognosis and immune infiltration patterns 5. Clinically, NSUN6 serves as a diagnostic biomarker in sepsis pathogenesis 6. Mutations in NSUN6 associate with autosomal recessive intellectual developmental disorder 82, highlighting its essential cellular role.