Abstract
The yeast Saccharomyces cerevisiae Crt1 transcription repressor is an effector of the DNA damage and replication checkpoint pathway. Crt1 binds and represses genes encoding ribonucleotide reductase (RNR) and its own promoter, establishing a negative-feedback pathway. The role of Rfx1, the mammalian Crt1 homologue, remained uncertain. In this study we investigated the possibility that Rfx1 plays a similar function in animal cells. We show here that, like Crt1, Rfx1 binds and represses its own promoter. Furthermore, Rfx1 binding to its promoter is reduced upon induction of a DNA replication block by hydroxyurea, which led to a release of repression. Significantly, like Crt1, Rfx1 binds and represses the RNR-R2 gene. Upon blocking replication and UV treatment, expression of both Rfx1 and RNR-R2 is induced; however, unlike the results seen with the RNR-R2 gene, the derepression of the RFX1 gene is only partially blocked by inhibiting Chk1, the DNA checkpoint kinase. This report provides evidence for a common mechanism for Crt1 and Rfx1 expression and for the conservation of their mode of action in response to a DNA replication block. We suggest that Rfx1 plays a role in the DNA damage response by down-regulating a subset of genes whose expression is increased in response to replication blocking and UV-induced DNA damage.
Highlights
Cells respond to DNA damage and replication blocking by attenuating cell cycle progression via transcriptional and posttranscriptional regulation of the components of the DNA replication, repair, and recombination pathways
Upon blocking replication and UV treatment, expression of both Rfx1 and ribonucleotide reductase (RNR)-R2 is induced; unlike the results seen with the RNR-R2 gene, the derepression of the RFX1 gene is only partially blocked by inhibiting Chk1, the DNA checkpoint kinase
We suggest that Rfx1 plays a role in the DNA damage response by down-regulating a subset of genes whose expression is increased in response to replication blocking and UV-induced DNA damage
Summary
Cells respond to DNA damage and replication blocking by attenuating cell cycle progression via transcriptional and posttranscriptional regulation of the components of the DNA replication, repair, and recombination pathways. In Saccharomyces cerevisiae, RNR inhibition decreases deoxynucleoside triphosphate (dNTP) levels and activates a DNA checkpoint pathway composed of the Mec, Rad, and Dun protein kinases [21]. It has been proposed that inhibition of an autoregulatory repressor in response to DNA damage is a strategy conserved throughout prokaryotic and eukaryotic evolution [21], but whether a similar Crt related mechanism exists in animal cells was not reported. The level of conservation raises the possibility that the Rfx family has retained its roles along the course of evolution Such a model is supported by the recent identification of Drosophila Rfx, dRfx, that is involved in cell cycle regulation [32] in addition to the previously described dRfx that is involved in cilium formation similar to that seen with C. elegans DAF-19 and the mammalian Rfx3 [6, 12, 39]
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