Article

A Screen for Suppressors of Gross Chromosomal Rearrangements Identifies a Conserved Role for PLP in Preventing DNA Lesions

Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada.
PLoS Genetics (Impact Factor: 8.17). 09/2007; 3(8):e134. DOI: 10.1371/journal.pgen.0030134
Source: PubMed

ABSTRACT Genome instability is a hallmark of cancer cells. One class of genome aberrations prevalent in tumor cells is termed gross chromosomal rearrangements (GCRs). GCRs comprise chromosome translocations, amplifications, inversions, deletion of whole chromosome arms, and interstitial deletions. Here, we report the results of a genome-wide screen in Saccharomyces cerevisiae aimed at identifying novel suppressors of GCR formation. The most potent novel GCR suppressor identified is BUD16, the gene coding for yeast pyridoxal kinase (Pdxk), a key enzyme in the metabolism of pyridoxal 5' phosphate (PLP), the biologically active form of vitamin B6. We show that Pdxk potently suppresses GCR events by curtailing the appearance of DNA lesions during the cell cycle. We also show that pharmacological inhibition of Pdxk in human cells leads to the production of DSBs and activation of the DNA damage checkpoint. Finally, our evidence suggests that PLP deficiency threatens genome integrity, most likely via its role in dTMP biosynthesis, as Pdxk-deficient cells accumulate uracil in their nuclear DNA and are sensitive to inhibition of ribonucleotide reductase. Since Pdxk links diet to genome stability, our work supports the hypothesis that dietary micronutrients reduce cancer risk by curtailing the accumulation of DNA damage and suggests that micronutrient depletion could be part of a defense mechanism against hyperproliferation.

0 Followers
 · 
108 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: Toxic DNA-protein crosslinks (DPCs) arise by ionizing irradiation and UV light, are particularly caused by endogenously produced reactive compounds such as formaldehyde, and also occur during compromised topoisomerase action. Although nucleotide excision repair and homologous recombination contribute to cell survival upon DPCs, hardly anything is known about mechanisms that target the protein component of DPCs directly. Here, we identify the metalloprotease Wss1 as being crucial for cell survival upon exposure to formaldehyde and topoisomerase 1-dependent DNA damage. Yeast mutants lacking Wss1 accumulate DPCs and exhibit gross chromosomal rearrangements. Notably, in vitro assays indicate that substrates such as topoisomerase 1 are processed by the metalloprotease directly and in a DNA-dependent manner. Thus, our data suggest that Wss1 contributes to survival of DPC-harboring cells by acting on DPCs proteolytically. We propose that DPC proteolysis enables repair of these unique lesions via downstream canonical DNA repair pathways.
    Cell 07/2014; 158(2). DOI:10.1016/j.cell.2014.04.053 · 33.12 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Pyridoxal 5'-phosphate (PLP), the active form of vitamin B6, has been implicated in preventing human pathologies, such as diabetes and cancer. However, the mechanisms underlying the beneficial effects of PLP are still unclear. Using Drosophila as a model system, we show that PLP deficiency, caused either by mutations in the pyridoxal kinase-coding gene (dPdxk) or by vitamin B6 antagonists, results in chromosome aberrations (CABs). The CAB frequency in PLP-depleted cells was strongly enhanced by sucrose, glucose or fructose treatments, and dPdxk mutant cells consistently displayed higher glucose contents than their wild type counterparts, an effect that is at least in part a consequence of an acquired insulin resistance. Together, our results indicate that a high intracellular level of glucose has a dramatic clastogenic effect if combined with PLP deficiency. This is likely due to an elevated level of Advanced Glycation End-products (AGE) formation. Treatment of dPdxk mutant cells with α-lipoic acid (ALA) lowered both AGE formation and CAB frequency, suggesting a possible AGE-CAB cause-effect relationship. The clastogenic effect of glucose in PLP-depleted cells is evolutionarily conserved. RNAi-mediated silencing of PDXK in human cells or treatments with PLP inhibitors resulted in chromosome breakage, which was potentiated by glucose and reduced by ALA. These results suggest that patients with concomitant hyperglycemia and vitamin B6 deficiency may suffer chromosome damage. This might impact cancer risk, as CABs are a well-known tumorigenic factor.
    PLoS Genetics 03/2014; 10(3):e1004199. DOI:10.1371/journal.pgen.1004199 · 8.17 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Whole genome sequencing of cancer genomes has revealed a diversity of recurrent gross chromosomal rearrangements (GCRs) that are likely signatures of specific defects in DNA damage response pathways. However, inferring the underlying defects has been difficult due to insufficient information relating defects in DNA metabolism to GCR signatures. By analyzing over 95 mutant strains of Saccharomyces cerevisiae, we found that the frequency of GCRs that deleted an internal CAN1/URA3 cassette on chrV L while retaining a chrV L telomeric hph marker was significantly higher in tel1Δ, sae2Δ, rad53Δ sml1Δ, and mrc1Δ tof1Δ mutants. The hph-retaining GCRs isolated from tel1Δ mutants contained either an interstitial deletion dependent on non-homologous end-joining or an inverted duplication that appeared to be initiated from a double strand break (DSB) on chrV L followed by hairpin formation, copying of chrV L from the DSB toward the centromere, and homologous recombination to capture the hph-containing end of chrV L. In contrast, hph-containing GCRs from other mutants were primarily interstitial deletions (mrc1Δ tof1Δ) or inverted duplications (sae2Δ and rad53Δ sml1Δ). Mutants with impaired de novo telomere addition had increased frequencies of hph-containing GCRs, whereas mutants with increased de novo telomere addition had decreased frequencies of hph-containing GCRs. Both types of hph-retaining GCRs occurred in wild-type strains, suggesting that the increased frequencies of hph retention were due to the relative efficiencies of competing DNA repair pathways. Interestingly, the inverted duplications observed here resemble common GCRs in metastatic pancreatic cancer.
    PLoS Genetics 04/2014; 10(4):e1004277. DOI:10.1371/journal.pgen.1004277 · 8.17 Impact Factor

Full-text (2 Sources)

Download
37 Downloads
Available from
Jun 2, 2014