The DNA replication machinery encounters problems at numerous genomic regions that are inherently difficult to replicate. These genomic regions include telomeres, which contain repetitive DNA and telomere-binding proteins. If not properly regulated, replication of such genomic regions can result in DNA damage, leading to genomic instability. Studies implicated a role of Timeless-related proteins at difficult-to-replicate genomic regions, including telomeres. However, how these proteins maintain telomeres was elusive. In a recent report, we described the role of Swi1, a Timeless-related protein, in telomere maintenance in fission yeast. We demonstrated that Swi1 is required for proper replication of repeat DNA sequences at telomeres. We also showed that Swi1-deficient cells utilize recombination-based ALT (alternative lengthening of telomeres)-like mechanisms to maintain telomeres in the absence of telomerase. Here, we highlight these findings and present additional data to discuss the role of Swi1Timeless in telomere protection and ALT prevention.

The Schizosaccharomyces pombe MBF complex activates the transcription of genes required for DNA synthesis and S phase. The MBF complex contains several proteins, including the core components Cdc10, Res1 and Res2, the co-repressor proteins Yox1 and Nrm1 and the co-activator Rep2. It has recently been shown how MBF is regulated when either the DNA damage or the DNA synthesis checkpoints are activated. However, how MBF is regulated in a normal unperturbed cell cycle is still not well understood. We have set up a genome-wide genomic screen searching for global regulators of MBF. We have crossed our knock-out collection library with a reporter strain that allows the measurement of MBF activity in live cells by flow cytometry. We confirm previously known regulators of MBF and show that COP9/ signalosome and tRNA methyltransferases also regulate MBF activity.

In fission yeast, MBF-dependent transcription is inactivated at the end of S phase through a negative feedback loop that involves the co-repressors, Yox1 and Nrm1. Although this repression system is well known, the molecular mechanisms involved in MBF activation remain largely unknown. Compacted chromatin constitutes a barrier to activators accessing promoters. Here, we show that chromatin regulation plays a key role in activating MBF-dependent transcription. Gcn5, a part of the SAGA complex, binds to MBF-regulated promoters through the MBF co-activator Rep2 in a cell cycle-dependent manner and in a reverse correlation to the binding of the MBF co-repressors, Nrm1 or Yox1. We propose that the co-repressors function as physical barriers to SAGA recruitment onto MBF promoters. We also show that Gcn5 acetylates specific lysine residues on histone H3 in a cell cycle-regulated manner. Furthermore, either in a gcn5 mutant or in a strain in which histone H3 is kept in an unacetylated form, MBF-dependent transcription is downregulated. In summary, Gcn5 is required for the full activation and correct timing of MBF-regulated gene transcription.

Chromatin structure is an essential factor in the proper regulation of DNA repair, DNA replication and transcription. The INO80 complex and the SWR complex have been shown to play a fundamental role in transcription regulation through remodeling chromatin at specific genes and loci. Here, we report that the Schizosaccharomyces pombe INO80 complex physically interacts with the mlui-binding factor (MBF) complex. Furthermore, we are able to detect the INO80 complex in MBF-regulated promoters. Binding of INO80 to these genes is cell cycle regulated, with a maximum binding preceding their transcription and accumulation of their mRNAs. In fact, the INO80 complex is required to fully and timely activate the transcription of these genes. We also show that the accumulation of acetylated H2A.Z at the +1 nucleosome is cell cycle regulated. Cells in which H2A.Z acetylation is abolished still have some cell cycle-regulated transcription of MBF-dependent genes, although to a much lesser extent.