Autorepression of Rfx1 Gene Expression: Functional Conservation from Yeast to Humans in Response to DNA Replication Arrest

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.

Hepatitis B Virus Activates Deoxynucleotide Synthesis in Nondividing Hepatocytes by Targeting the R2 Gene

“…The R2 gene is under repression by the Rfx1 transcription factor. 11 This mechanism of R2 repression was first identified in yeast where the Rfx1 ortholog, Crt1, represses the R2 gene. 29 Consistent with this repression role of Rfx1, chromatin immunoprecipitation (ChIP) analysis revealed that Rfx1 was bound to the R2 promoter more effectively in the quiescent cells than in the proliferative cells (Fig.…”

“…Chromatin immunoprecipitation (ChIP) was performed as described. 11 Western Blotting. Western blotting (protein immunoblotting) was performed as previously described.…”

“…Western blotting (protein immunoblotting) was performed as previously described. 11,13 Proteins were detected with the following antibodies: goat anti-R2 (N-18: SC-10844; Santa Cruz Biotechnology, Santa Cruz, CA), mouse antibody to hepatitis B core antigen (anti-HBcAg), 13 and rabbit anti-Rfx1 polyclonal antibodies. 11 The blots were then reacted with horseradish peroxidase-conjugated secondary antibody (Jackson), and enhanced chemiluminescence (ECL) detection was performed using EZ-ECL (Biological Industries).…”

“…We further show that hepatitis B x protein (HBx), a regulatory protein of HBV, is sufficient for R2 induction by blocking the access of regulatory factor x1 (Rfx1), a repressor of the R2 gene. 11 …”

DNA damage-induced upregulation of miR-709 in the germline downregulates BORIS to counteract aberrant DNA hypomethylation

“…Interestingly, Rfx1 has been implicated in DNA damage recognition, and it is also a downstream target of the DNA damage sensor ATR kinase. 20 Rfx1 upregulation might be as follows: ATR is induced by DNA damage caused by IR, then ATR acts on its downstream targets including Rfx1. Upregulation of Rfx1 consequentially upregulates miR-709, which then inhibits the translation of BORIS and affects DNA methylation process in the germline (Fig.…”