The ATM repair pathway inhibits RNA polymerase I transcription in response to chromosome breaks

Experimental Immunology, NCI.
Nature (Impact Factor: 41.46). 07/2007; 447(7145):730-4. DOI: 10.1038/nature05842
Source: PubMed


DNA lesions interfere with DNA and RNA polymerase activity. Cyclobutane pyrimidine dimers and photoproducts generated by ultraviolet irradiation cause stalling of RNA polymerase II, activation of transcription-coupled repair enzymes, and inhibition of RNA synthesis. During the S phase of the cell cycle, collision of replication forks with damaged DNA blocks ongoing DNA replication while also triggering a biochemical signal that suppresses the firing of distant origins of replication. Whether the transcription machinery is affected by the presence of DNA double-strand breaks remains a long-standing question. Here we monitor RNA polymerase I (Pol I) activity in mouse cells exposed to genotoxic stress and show that induction of DNA breaks leads to a transient repression in Pol I transcription. Surprisingly, we find Pol I inhibition is not itself the direct result of DNA damage but is mediated by ATM kinase activity and the repair factor proteins NBS1 (also known as NLRP2) and MDC1. Using live-cell imaging, laser micro-irradiation, and photobleaching technology we demonstrate that DNA lesions interfere with Pol I initiation complex assembly and lead to a premature displacement of elongating holoenzymes from ribosomal DNA. Our data reveal a novel ATM/NBS1/MDC1-dependent pathway that shuts down ribosomal gene transcription in response to chromosome breaks.

Download full-text


Available from: Michael J Kruhlak, May 20, 2015
  • Source
    • "requires ATM in mammalian cells (Kruhlak et al., 2007; 10.3389/fgene.2013.00136/full, "
    [Show abstract] [Hide abstract]
    ABSTRACT: Emerging evidence indicate that the mammalian checkpoint kinase ATM induces transcriptional silencing in cis to DNA double-strand breaks (DSBs) through a poorly understood mechanism. Here we show that in Saccharomyces cerevisiae a single DSB causes transcriptional inhibition of proximal genes independently of Tel1/ATM and Mec1/ATR. Since the DSB ends undergo nucleolytic degradation (resection) of their 5'-ending strands, we investigated the contribution of resection in this DSB-induced transcriptional inhibition. We discovered that resection-defective mutants fail to stop transcription around a DSB, and the extent of this failure correlates with the severity of the resection defect. Furthermore, Rad9 and generation of γH2A reduce this DSB-induced transcriptional inhibition by counteracting DSB resection. Therefore, the conversion of the DSB ends from double-stranded to single-stranded DNA, which is necessary to initiate DSB repair by homologous recombination, is responsible for loss of transcription around a DSB in S. cerevisiae.
    eLife Sciences 07/2015; 4. DOI:10.7554/eLife.08942 · 9.32 Impact Factor
  • Source
    • "By combining these results with our recent work on the genomic architecture of NORs, we can now present a model for how integrity of rDNA arrays is maintained against an onslaught of DNA damage (Fig. 7). While the use of γ-irradiation and laser microirradiation to induce DSBs has yielded conflicting results (Kruhlak et al. 2007; Moore et al. 2011; Larsen et al. 2014), here we unequivocally demonstrated that DSBs within rDNA are sufficient to induce ATM-dependent inhibition of Pol I transcription. The introduction of the CRISPR/Cas9 allowed us to demonstrate that DSBs in both transcribed and nontranscribed regions of the rDNA repeat induce this response. "
    [Show abstract] [Hide abstract]
    ABSTRACT: DNA double-strand breaks (DSBs) are repaired by two main pathways: nonhomologous end-joining and homologous recombination (HR). Repair pathway choice is thought to be determined by cell cycle timing and chromatin context. Nucleoli, prominent nuclear subdomains and sites of ribosome biogenesis, form around nucleolar organizer regions (NORs) that contain rDNA arrays located on human acrocentric chromosome p-arms. Actively transcribed rDNA repeats are positioned within the interior of the nucleolus, whereas sequences proximal and distal to NORs are packaged as heterochromatin located at the nucleolar periphery. NORs provide an opportunity to investigate the DSB response at highly transcribed, repetitive, and essential loci. Targeted introduction of DSBs into rDNA, but not abutting sequences, results in ATM-dependent inhibition of their transcription by RNA polymerase I. This is coupled with movement of rDNA from the nucleolar interior to anchoring points at the periphery. Reorganization renders rDNA accessible to repair factors normally excluded from nucleoli. Importantly, DSBs within rDNA recruit the HR machinery throughout the cell cycle. Additionally, unscheduled DNA synthesis, consistent with HR at damaged NORs, can be observed in G1 cells. These results suggest that HR can be templated in cis and suggest a role for chromosomal context in the maintenance of NOR genomic stability. © 2015 van Sluis and McStay; Published by Cold Spring Harbor Laboratory Press.
    Genes & Development 06/2015; 29(11). DOI:10.1101/gad.260703.115 · 10.80 Impact Factor
  • Source
    • "Interestingly, Treacle, like MDC1, contains several phoshorylated SDT domains that mediate NBS1 binding, suggesting a conserved mechanism of NBS1 retention in the nucleus and nucleolus. It is notable that while MDC1 is not required for NBS1 retention to nucleoli, MDC1 deficient cells failed to silence rDNA transcription after IR treatment [57]. This may reflect a difference between the techniques used, as IR could induce damage directly in nucleolar DNA, or could indicate that MRE11 complex localization per se is not sufficient for rDNA silencing. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Newly identified interactions with the FHA and BRCT domains of NBS1 influence subcellular localization of the MRE11 complex.•The MRE11 complex promotes TOPBP1 recruitment and ATR activation during replication stress.•The MRE11 complex is a barrier to oncogene-induced tumorigenesis.
    Experimental Cell Research 10/2014; 329(1). DOI:10.1016/j.yexcr.2014.10.010 · 3.25 Impact Factor
Show more