Supplementary Table 1. List of DNA probes in ligation assay. Supplementary Table 2. List of DNA probes in EMSA assay. Supplementary Table 3. miRNAs identified by HTS that could induce excess DNA replication (Cut-off {greater than or equal to}25%). Supplementary Table 4. Prediction of seed-binding sites in miR-571 and 3'-UTR region (88-94) of GMNN (Geminin) based on Targetscan software. Supplementary Table 5. List of Major DNA replication and cell cycle related genes (n=105)
ARTICLE ABSTRACT
DNA rereplication leads to genomic instability and has been implicated in the pathology of a variety of human cancers. Eukaryotic DNA replication is tightly controlled to ensure it occurs only once during each cell cycle. Geminin is a critical component of this control, it prevents DNA rereplication from occurring during S, G2, and early M phases by preventing MCM helicases from forming prereplication complexes. Geminin is targeted for degradation by the anaphase-promoting complex (APC/C) from anaphase through G1-phase, however, accumulating evidence indicates that Geminin is downregulated in late S-phase due to an unknown mechanism. Here, we used a high-throughput screen to identify miRNAs that can induce excess DNA replication and found that miR-571 could reduce the protein level of Geminin in late S-phase independent of the APC/C. Furthermore, miR-571 regulated efficient DNA replication and S-phase cell-cycle progression. Strikingly, c-Myc suppressed miR-571 expression by binding directly to the miR-571 promoter. At the beginning of S-phase, Cdk2 phosphorylated c-Myc at Serine 62, promoting its association with the miR-571 promoter region. Collectively, we identify miR-571 as the first miRNA that prevents aberrant DNA replication and the Cdk2–c-Myc–miR-571 axis as a new pathway for regulating DNA replication, cell cycle, and genomic stability in cancer cells.
These findings identify a novel regulatory mechanism that is critical for maintaining genome integrity by regulating DNA replication and cell-cycle progression.
