Tumor suppressor p53 plays a central role in the DNA damage response. DNA-damaging agents modulate nuclear actin dynamics, influencing cell behaviors; however, it remains unclear whether p53 affects the structure of nuclear actin filaments. Here, we show that actin filament formation in the nucleus induced by the DNA-damaging agent doxorubicin (DOXO) treatment is suppressed by p53 via caspase-1 expression. In DOXO-treated cells, the nuclear actin filament formation was promoted by p53 depletion. Whilst the DOXO treatment caused an increase in caspase-1 expression, which was largely attenuated by p53 depletion, overexpression of caspase-1 reduced DOXO-induced formation of nuclear actin filaments in p53-depleted cells. By contrast, caspase-1 inhibition, together with DOXO treatment, induced nuclear actin filament formation even in cells bearing wild-type p53. These results suggest that p53-mediated expression of caspase-1 suppresses nuclear actin filament formation. In addition, we found that expression of nLifeact-GFP, which contains the filamentous actin (F-actin)-binding peptide and nuclear localization signal (NLS), promoted the bundling of nuclear actin filaments and strongly altered the chromatin structure in DOXO-treated, p53-depleted cells. Given that the loss of p53 is associated with cancer progression, the results of this study raised the possibility that artificial reinforcement of nuclear actin filaments by nLifeact-GFP may enhance the cytotoxicity effect of DNA-damaging agents in aggressive cancer cells.