Mind the gap: keeping UV lesions in check

Abstract

Cells respond to genotoxic insults by triggering a DNA damage checkpoint surveillance mechanism and by activating repair pathways. Recent findings indicate that the two processes are more related than originally thought. Here we discuss the mechanisms involved in responding to UV-induced lesions in different phases of the cell cycle and summarize the most recent data in a model where Nucleotide Excision Repair (NER) and exonucleolytic activities act in sequence leading to checkpoint activation in non replicating cells. The critical trigger is likely represented by problematic intermediates that cannot be completely or efficiently repaired by NER. In S phase cells, on the other hand, the replicative polymerases, blocked by bulky UV lesions, re-initiate DNA synthesis downstream of the lesions, leaving behind a ssDNA tract. If these gaps are not rapidly refilled, checkpoint kinases will be activated.

Fig. 1

The DNA damage checkpoint cascade. The DNA damage checkpoint is triggered by a ssDNA region. The left side of the figure reports the checkpoint cascade in budding yeast. RPA-covered ssDNA recruits the Mec1-Ddc2 and the 9-1-1 complexes. Phosphorylated Ddc1 interacts with Dpb11 which recruits the Rad9 mediator. Rad53 and Chk1 kinases are activated upon binding to oligomeric Rad9 and then leave chromatin to find their own targets. The right side of the figure summarizes the same signaling cascade in human cells.

Fig. 2

UV-induced checkpoint response. In cells that are not replicating their genome (i.e., G1, G2 or non-cycling cells), NER removes UV lesions efficiently and DNA polymerases (i.e., pol δ, pol ɛ, TLS polymerases) begin the refilling process. If the repair reaction is impeded after the excision step, a competition between the refilling polymerases and Exo1 nuclease can take place. Problematic refilling (e.g., closely opposing lesions) allows Exo1 to further process the gapped intermediate generating long ssDNA gaps which recruit checkpoint factors and trigger the signaling. At low UV-doses G1 and G2 cells do not accumulate large ssDNA gaps since UV lesions can be efficiently removed by NER. If the damages are still present when the cell enters S phase, the replicative polymerase will be blocked by the bulky lesion and will reinitiate DNA synthesis further downstream, leaving ssDNA gaps behind the replication fork. These gaps can be refilled by post replication repair and trigger a post-replication checkpoint.

Fig. 3

Crosstalks between NER and checkpoint factors. A two-way functional interaction exists between the checkpoint machinery and the NER apparatus: some examples (discussed in the text) are shown. (A) NER factors recruit checkpoint proteins to damaged chromosomes, thus facilitating the activation of the signaling cascade. (B) DNA damage checkpoint factors modulate NER activity allowing for efficient repair of the lesions.