DNA Topoisomerase III Alpha Regulates p53-Mediated Tumor Suppression

Purpose: Human DNA topoisomerase III alpha (hTOP3α) is involved in DNA repair surveillance and cell-cycle checkpoints possibly through formatting complex with tumor suppressors. However, its role in cancer development remained unsolved. Experimental Design: Coimmunoprecipitation, sucrose gradient, chromatin immunoprecipitation (ChIP), real time PCR, and immunoblotting analyses were performed to determine interactions of hTOP3α with p53. Paired cell lines with different hTOP3α levels were generated via ectopic expression and short hairpin RNA (shRNA)-mediated knockdown approaches. Cellular tumorigenic properties were analyzed using cell counting, colony formation, senescence, soft agar assays, and mouse xenograft models. Results: The hTOP3α isozyme binds to p53 and cofractionizes with p53 in gradients differing from fractions containing hTOP3α and BLM. Knockdown of hTOP3α expression (sh-hTOP3α) caused a higher anchorage-independent growth of nontumorigenic RHEK-1 cells. Similarly, sh-hTOP3α and ectopic expression of hTOP3α in cancer cell lines caused increased and reduced tumorigenic abilities, respectively. Genetic and mutation experiments revealed that functional hTOP3α, p53, and p21 are required for this tumor-suppressive activity. Mechanism-wise, ChIP data revealed that hTOP3α binds to the p53 and p21 promoters and positively regulates their expression. Two proteins affect promoter recruitments of each other and collaborate in p21 expression. Moreover, sh-hTOP3α and sh-p53 in AGS cells caused a similar reduction in senescence and hTOP3α mRNA levels were lower in gastric and renal tumor samples. Conclusion: We concluded that hTOP3α interacts with p53, regulates p53 and p21 expression, and contributes to the p53-mediated tumor suppression. Clin Cancer Res; 20(6); 1489–501. ©2014 AACR.