NOP53 undergoes liquid-liquid phase separation and promotes tumor radio-resistance.

Aberrant DNA damage response (DDR) axis remains the major molecular mechanism for tumor radio-resistance. We recently characterized liquid-liquid phase separation (LLPS) as an essential mechanism of DDR, and identified several key DDR factors as potential LLPS proteins, including nucleolar protein NOP53. In this study, we found that NOP53 formed highly concentrated droplets in vivo and in vitro, which had liquid-like properties including the fusion of adjacent condensates, rapid fluorescence recovery after photobleaching and the sensitivity to 1,6-hexanediol. Moreover, the intrinsically disordered region 1 (IDR1) is required for NOP53 phase separation. In addition, multivalent-arginine-rich linear motifs (M-R motifs), which are enriched in NOP53, were essential for its nucleolar localization, but were dispensable for the LLPS of NOP53. Functionally, NOP53 silencing diminished tumor cell growth, and significantly sensitized colorectal cancer (CRC) cells to radiotherapy. Mechanically, NOP53 negatively regulated p53 pathway in CRC cells treated with or without radiation. Importantly, data from clinical samples confirmed a correlation between NOP53 expression and tumor radio-resistance. Together, these results indicate an important role of NOP53 in radio-resistance, and provide a potential target for tumor radio-sensitization.