TY  - GEN
T1  - Top1p targeting by Fob1p at the ribosomal Replication Fork Barrier does not account for camptothecin sensitivity in Saccharomyces cerevisiae cells
AU  - Pourali, Pardis
AU  - Pasero, Philippe
AU  - Pardo, Benjamin
DO  - 10.17912/micropub.biology.000514
UR  - http://beta.micropublication.org/journals/biology/micropub-biology-000514/
AB  - DNA topoisomerases play a vital role in solving topological constrains during DNA replication and transcription (Pommier et al. 2016). Top1p is a DNA topoisomerase that relaxes DNA supercoiling by nicking the DNA, creating a covalent bond between the enzyme and the 3’ end of the DNA, called Top1p cleavage complex (Top1cc). Once the DNA is relaxed, Top1p religates the break by reversing its covalent binding. Top1p activity can be inhibited by drugs such as camptothecin (CPT), whose derivatives (topotecan, irinotecan) are largely used in cancer therapy. CPT binds to the Top1cc and delay the religation reaction, thus blocking Top1p on DNA (Figure 1A) (Pommier et al. 2006). It has been shown that treatment of cells with CPT induces DNA double-strand breaks (DSBs) specifically during DNA replication. One model proposes that the DNA nick is converted into a DSB by the passage of the replication fork, which separates the parental duplex DNA (Strumberg et al. 2000). Because CPT prevents Top1p ability to remove topological stress (Koster et al. 2007), another model proposed that the accumulation of topological constrains would block replication fork advance and induce DSB formation as a consequence of fork cleavage by the MUS81 nuclease (Regairaz et al. 2011). These DSBs must be repaired, otherwise mutations could arise and affect genome stability. DSBs are mainly repaired by Homologous Recombination (HR) during the S and G2 phases of the cell cycle, the Rad52p protein being a key player in this pathway in yeast Saccharomyces cerevisiae (Pardo et al. 2009). Rad52p proteins relocalize into discrete sub nuclear foci upon DSB formation (Lisby et al. 2003). In response to CPT treatment, yeast cells accumulate Rad52p foci both in the nucleus and the nucleolus (Stuckey et al. 2015). These results suggest that CPT-induced DSBs arise in the ribosomal DNA (rDNA) genes, which reside in the nucleolus. These genes are organized on the chromosome XII in a single cluster of 150-200 tandem repeats (Kobayashi 2011). Each rDNA repeat includes a Replication Fork Barrier (rRFB) bound by Fob1p, whose function is to avoid collisions between replication and transcription machineries by stalling the movement of replication forks in only one direction (Kobayashi and Horiuchi 1996). In the rDNA, it has been shown that Top1ccs accumulate naturally and specifically at the rRFB, and that this accumulation is completely lost in the absence of Fob1p (Krawczyk et al. 2014). DSBs have also been detected at the rRFB and are dependent on Top1p and Fob1p. These DSB signals were increased upon the inhibition of Top1p by CPT (Krawczyk et al. 2014), suggesting that the accumulation of Fob1p-dependent Top1ccs at the rRFB leads to DSB formation. Since the rDNA represents about 10% of the genome size of Saccharomyces cerevisiae and these DSBs could be detrimental for cell survival, we thus investigated if the absence of Fob1p could improve the resistance of yeast cells to CPT.
PY  - 2022
JO  - microPublication Biology
ER  -