NAT10 maintains stem cell homeostasis by mitigating mRNA decay through an ac4C-independent mechanism.

Haematopoietic stem cells (HSCs) represent a well-established system for studying stem cell maintenance. While RNA regulators have been reported in HSCs, a systematic characterization and how they define transcript fate remains outstanding. Here we profile RNA characteristics of HSC-essential genes and uncover a notable feature in both human and mouse: they have extended 3' untranslated regions specifically enriched with AU-rich elements (AREs). These AREs are crucial for the expression of HSC genes, primarily through NAT10, which stabilizes their mRNAs. Notably, Nat10 deficiency markedly disrupts HSCs self-renewal and long-term reconstitution capacity. Mechanistically, NAT10 recruits ribosomes to the 3' untranslated region AREs of HSC-essential mRNAs, sheltering them from degradation-an effect independent of NAT10's ac4C catalytic activity. Moreover, NAT10 dysregulations were associated with multiple human haematological malignancies. Collectively, our findings uncover a specific mechanism of RNA turnover control mediated by specific RNA ARE motifs and identify a non-catalytic role of NAT10 in maintaining HSC homeostasis.