Top: During DNA replication stress, cells accumulate ssDNA that is repaired by mutagenic TLS polymerases.
Bottom: Single molecule DNA analysis shows that ssDNA (shown in red) accumulates on DNA (shown in green) when RAD5 is absent.

Grant Brown’s lab at the Donnelly Centre has characterized a new mutagenic ssDNA gap filling pathway in budding yeast. Their findings will be published in the March 7th issue of Molecular Cell.

DNA replication is a fundamental process that occurs before every cell division and must be completed in a timely and accurate fashion. Under normal cellular conditions this reaction is carried out by the highly processive and accurate replicative polymerases. However, DNA replication is often stressed and impeded by physical lesions on DNA, tightly bound protein or RNA complexes, deprivation of nucleotides or mutation of the DNA replication machinery – collectively referred to as DNA replication stress. Cells respond to replication stress by promoting completion of bulk DNA replication during S-phase, leaving ssDNA gaps behind for small-scale repair before cell division

The current study, spearheaded by Dave Gallo and colleagues, identified a new mode of mutagenic gap filling mediated by Trans-lesion synthesis (TLS) polymerases. TLS polymerases have a larger active site to accommodate non-canonical DNA templates and lack-proof reading activity, making them error-prone. This led to the belief they are only utilized when the replication fork encounters a DNA damage lesion during DNA replication. The finding that TLS polymerases are active on undamaged templates was surprising and points to an expanded utilization of TLS polymerases in replicating eukaryotic genomes.

But why would cells choose a mutagenic pathway to fill ssDNA gaps? And does this occur in human cells? These questions remain to be answered.

We know that when cells fail to repair ssDNA gaps before cell division there is increased chromosome rearrangements and breakage leading to genome catastrophe and cell death. We also know that DNA replication stress is a hallmark of cancer. Expression of certain oncogenes leads to deregulated DNA replication dynamics in a process called oncogene-induced replication stress. Cells experiencing oncogene-induced replication stress upregulate the DNA replication stress response to allow continued growth and cell division. Inhibiting the DNA replication stress response in cancer cells offers a selective way to kill cancer cells, leaving healthy cells unaffected. The finest example of this is PARP inhibitors, currently being used to treat cancers. It is possible that, like budding yeast, cancer cells also upregulate TLS polymerases to cope with replication stress, offering a new pathway to target with therapeutics to treat cancer.

This research was supported by the Canadian Cancer Society and the Natural Sciences and Engineering Research Council of Canada.