Linking DNA replication checkpoint to MBF cell-cycle transcription reveals a distinct class of G1/S genes

Abstract

Reprogramming gene expression is crucial for DNA replication stress response. We used quantitative proteomics to establish how the transcriptional response results in changes in protein levels. We found that expression of G1/S cell-cycle targets are strongly up-regulated upon replication stress, and show that MBF, but not SBF genes, are up-regulated via Rad53-dependent inactivation of the MBF co-repressor Nrm1. A subset of G1/S genes was found to undergo an SBF-to-MBF switch at the G1/S transition, enabling replication stress-induced transcription of genes targeted by SBF during G1. This subset of G1/S genes is characterized by an overlapping Swi4/ Mbp1-binding site and is enriched for genes that cause a cell cycle and/or growth defect when overexpressed. Analysis of the prototypical switch gene TOS4 (Target Of SBF 4) reveals its role as a checkpoint effector supporting the importance of this distinct class of G1/S genes for the DNA replication checkpoint response. Our results reveal that replication stress induces expression of G1/S genes via the Rad53-MBF pathway and that an SBF-to-MBF switch characterizes a new class of genes that can be induced by replication stress.

The checkpoint transcriptional response: Make sure to turn it off once you are satisfied

“…In both cases, transcriptional activation is mediated via checkpointdependent inhibition of a transcriptional repressor (Crt1 for DDR and Nrm1 for CC) that participates in negative feedback regulation. 13,14 This repressor-mediated regulation enables transcription to be rapidly repressed once cells have dealt Consistent with the notion that the DNA damage checkpoint is a central trigger for most of the transcriptional responses to replication stress, the vast majority of the observed replication-specific changes in expression were indeed found to be dependent on the Mec1/Rad53 kinasesignaling pathway. 47,49 This pathway can be divided into two major branches, one mediated by the most downstream checkpoint protein kinase Dun1, a kinase that is directly phosphorylated and activated by Rad53, while the other branch is Dun1-independent (Fig.…”

“…13,51 Under unperturbed conditions, replication stress response is not fully understood. 27 Budding yeast is a very useful system to study the replication checkpoint due to a high degree of conservation of the main checkpoint kinases.…”

“…This group of replication stress-induced G 1 /S genes includes close to 100 genes and consequently represents one of the largest single groups of co-regulated targets among DNA-damage induced genes. 13,51 It is well established that G 1 /S transcription depends on the SBF and MBF transcription factor complexes that regulate over 200 cell cycle genes. 53 Our recent work, published in The EMBO Journal, revealed that CC genes that depend on MBF for inactivation once cells progress into S phase are transcriptionally induced upon replication stress.…”

“…65,66 Moreover, analysis of transcription induction upon replication stress showed that this group of genes is enriched for genes that are upregulated during replication stress conditions. 13,51 Switch Genes as DosageSensitive Checkpoint Effectors TOS genes. Among the genes found to be regulated by a SBF-to-MBF switch, seven were previously identified in the Iyer ChIP-chip study as targets of SBF (TOS1, 2, 3, 4, 6 and 8).…”

“…Interestingly, we found that this set of genes included not only G 1 /S genes solely regulated by MBF throughout the cell cycle, but also a new subset of G 1 /S genes regulated by SBF during G 1 and by MBF during S-phase. 13 This subset of genes was named "switch genes" based on the SBF-to-MBF switch at their promoters during the G 1 -to-S transition. The SBF-to-MBF switch was first established for the G 1 /S gene TOS4.…”