Transcription termination factor reb1p causes two replication fork barriers at its cognate sites in fission yeast rDNA in vivo

Abstract

Polar replication fork barriers (RFBs) near the 3'-end of the rRNA transcriptional unit are a conserved feature of rDNA replication in eukaryotes. In the mouse, in vivo studies indicate that the cis-acting Sal boxes required for rRNA transcription termination are also involved in replication fork blockage. On the contrary, in the budding yeast Saccharomyces cerevisiae, the rRNA transcription termination factors are not required for RFBs. Here we characterized the rDNA RFBs in the fission yeast Schizosaccharomyces pombe. S. pombe rDNA contains three closely spaced polar replication barriers named RFB1-3, in the 3’ to 5’ order. The transcription termination protein reb1 and its two binding sites, present at the 3’ end of the coding region, were required for fork arrest at RFB2 and RFB3 in vivo. On the other hand, fork arrest at the strongest RFB1 barrier was independent of the above transcription termination factors. Therefore, RFB2 and RFB3 resemble the barriers present in the mouse rDNA, whereas RFB1 is similar to the budding yeast RFBs. These results suggest that during evolution, cis- and trans-acting factors required for rRNA transcription termination became also involved in replication fork blockage. S. pombe would be a transitional species where both mechanisms coexist.