The Role of Transcription in Replication Blockage at Common Fragile Sites

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My previous studies suggested a relation between genomic regions containing very large transcriptionally active genes and preferred sites of disturbed replication, seen in metaphase chromosomes as uncondensed chromatin at common fragile sites (CFSs). CFSs are predetermined chromosomal breakage regions and have been implicated to have a causative role in cancer. The present proposal is aimed to study the interdependence of transcription, replication and chromosome condensation at CFSs. Some first results that I obtained by comparing different cell types in which three long genes are expressed or not, show indeed the involvement of transcription of these genes in CFS formation. I will now investigate if the colocalization of RNA Pol II with DNA polymerase is favoured at CFSs. In parallel, results of replication timing analysis across large CFS genes will be compared to active/inactive transcription and to CFS formation. By overexpressing positive transcription elongation factors I will investigate whether an abnormally slow transcription of these large genes could be a potential cause of the observed delayed replication at CFSs. To test for an involvement of perturbed coupling of transcription and replication in chromatin remodeling, I also plan to analyze histone modifications within CFSs. The combined results of this project are expected to (i) give new insights into the intertwining of distinct processes, such as Pol II transcription elongation, replication and chromosome condensation, in living cells and (ii) to better understand CFS formation.