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.

Download full-text


Available from: Diane C Cabelof, Aug 10, 2015
  • Source
    • "While microorganisms and plants have the ability to make the compound de novo, it is an essential nutrient in the diet of humans, being derived primarily from plant sources. Indeed, deficiency of the vitamin in humans and its association with various medical ailments has been well documented (Grillo et al., 2001; Stitt et al., 2002; Komatsu et al., 2003; Metz et al., 2003; Kanellis et al., 2007). All organisms have the ability to interconvert the different vitamer forms through a so-called salvage pathway (Figure 1). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Vitamin B₆ is an essential nutrient in the human diet derived primarily from plant sources. While it is well established as a cofactor for numerous metabolic enzymes, more recently, vitamin B₆ has been implicated as a potent antioxidant. The de novo vitamin B₆ biosynthesis pathway in plants has recently been unraveled and involves only two proteins, PDX1 and PDX2. To provide more insight into the effect of the compound on plant development and its role as an antioxidant, we have overexpressed the PDX proteins in Arabidopsis, generating lines with considerably higher levels of the vitamin in comparison with other recent attempts to achieve this goal. Interestingly, it was possible to increase the level of only one of the two catalytically active PDX1 proteins at the protein level, providing insight into the mechanism of vitamin B₆ homeostasis in planta. Vitamin B₆ enhanced lines have considerably larger vegetative and floral organs and although delayed in pre-reproductive development, do not have an altered overall morphology. The vitamin was observed to accumulate in seeds and the enhancement of its levels was correlated with an increase in their size and weight. This phenotype is predominantly a consequence of embryo enlargement as reflected by larger cells. Furthermore, plants that overaccumulate the vitamin have an increased tolerance to oxidative stress providing in vivo evidence for the antioxidant functionality of vitamin B₆. In particular, the plants show an increased resistance to paraquat and photoinhibition, and they attenuate the cell death response observed in the conditional flu mutant.
    The Plant Journal 05/2011; 66(3):414-32. DOI:10.1111/j.1365-313X.2011.04499.x · 6.82 Impact Factor
  • Source
    • "The GCR rate was measured on the basis of the previously reported protocol (Kanellis et al. 2007). Ten colonies from each strain were tested, and two rounds of independent experiments were conducted. "
    [Show abstract] [Hide abstract]
    ABSTRACT: This study reports an unusual ploidy-specific response to replication stress presented by a defective minichromosome maintenance (MCM) helicase allele in yeast. The corresponding mouse allele, Mcm4(Chaos3), predisposes mice to mammary gland tumors. While mcm4(Chaos3) causes replication stress in both haploid and diploid yeast, only diploid mutants exhibit G2/M delay, severe genetic instability (GIN), and reduced viability. These different outcomes are associated with distinct repair pathways adopted in haploid and diploid mutants. Haploid mutants use the Rad6-dependent pathways that resume stalled forks, whereas the diploid mutants use the Rad52- and MRX-dependent pathways that repair double strand breaks. The repair pathway choice is irreversible and not regulated by the availability of repair enzymes. This ploidy effect is independent of mating type heterozygosity and not further enhanced by increasing ploidy. In summary, a defective MCM helicase causes GIN only in particular cell types. In response to replication stress, early events associated with ploidy dictate the repair pathway choice. This study uncovers a fundamental difference between haplophase and diplophase in the maintenance of genome integrity.
    Genetics 04/2011; 187(4):1031-40. DOI:10.1534/genetics.110.125450 · 4.87 Impact Factor
  • Source
    • "In addition iron regulon genes are not induced upon benomyl treatment (Figure 4B, File S1, and Lucau- Danila et al. 2005). It is also important to note that, except for AFT1, no other iron regulon genes have been identified in genome-wide screens measuring genome instability by various assays (Kanellis et al. 2007; Yuen et al. 2007; Andersen et al. 2008). Our results indicate that the role of Aft1 in chromosome stability is distinct from its role as a transcriptional inducer of the iron regulon and iron homeostasis. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The Saccharomyces cerevisiae transcription factor Aft1 is activated in iron-deficient cells to induce the expression of iron regulon genes, which coordinate the increase of iron uptake and remodel cellular metabolism to survive low-iron conditions. In addition, Aft1 has been implicated in numerous cellular processes including cell-cycle progression and chromosome stability; however, it is unclear if all cellular effects of Aft1 are mediated through iron homeostasis. To further investigate the cellular processes affected by Aft1, we identified >70 deletion mutants that are sensitive to perturbations in AFT1 levels using genome-wide synthetic lethal and synthetic dosage lethal screens. Our genetic network reveals that Aft1 affects a diverse range of cellular processes, including the RIM101 pH pathway, cell-wall stability, DNA damage, protein transport, chromosome stability, and mitochondrial function. Surprisingly, only a subset of mutants identified are sensitive to extracellular iron fluctuations or display genetic interactions with mutants of iron regulon genes AFT2 or FET3. We demonstrate that Aft1 works in parallel with the RIM101 pH pathway and the role of Aft1 in DNA damage repair is mediated by iron. In contrast, through both directed studies and microarray transcriptional profiling, we show that the role of Aft1 in chromosome maintenance and benomyl resistance is independent of its iron regulatory role, potentially through a nontranscriptional mechanism.
    Genetics 05/2010; 185(3):1111-28. DOI:10.1534/genetics.110.117531 · 4.87 Impact Factor
Show more