Periodic Reporting for period 3 - REPLISTRESS (DNA Replication: From Physiology to Replication Stress in Human Cancer)
Période du rapport: 2021-10-01 au 2023-03-31
Aim 1. Determine whether origin firing in human cells is stochastic or deterministic; map the origin firing sites at high resolution; and identify functionally important origin motifs.
We have already mapped origin firing sites in human cells, genome-wide, at a resolution of 1 kb, which is the maximum resolution we can achieve by our EdUseq method. We have identified one motif that is present close to the origin firing sites and that is absolutely essential for origin firing and we are in the process of identifying additional motifs.
Aim 2. Determine the mechanisms by which transcription in G1 suppresses firing of intragenic origins.
We are pursuing two mechanisms. Either the transcription machinery displaces loaded MCMs or it modifies the chromatin in a way that prevents origin firing. We are currently working to distinguish between these two possibilities.
Aim 3. Determine why forks from intergenic origins do not collapse when replicating highly transcribed genes.
We studied origin firing and nascent transcription in normal mouse cells and have observed that most transcribed genes are replicated from origins that are upstream of the gene. This ensures that DNA replication and transcription are codirectional. This could explain why forks from intergenic origins do not collapse, when replicating transcribed genes.