Nicotinamide induces Fob1-dependent plasmid integration into chromosome XII in Saccharomyces cerevisiae.
In the ribosomal DNA (rDNA) array of Saccharomyces cerevisiae, DNA replication is arrested by the Fob1 protein in a site-specific manner that stimulates homologous recombination. The silent information regulator Sir2, which is loaded at the replication arrest sites by Fob1, suppresses this recombination event. A plasmid containing Fob1-binding sites, when propagated in a yeast strain lacking SIR2 is integrated into the yeast chromosome in a FOB1-dependent manner. We show that addition of nicotinamide (NAM) to the culture medium can stimulate such plasmid integration in the presence of SIR2. Pulsed-field gel electrophoresis analysis showed that plasmid integration occurred into chromosome XII. NAM-induced plasmid integration was dependent on FOB1 and on the homologous recombination gene RAD52. As NAM inhibits several sirtuins, we examined plasmid integration in yeast strains containing deletions of various sirtuin genes and observed that plasmid integration occurred only in the absence of SIR2, but not in the absence of other histone deacetylases. In the absence of PNC1 that metabolizes NAM, a reduced concentration of NAM was required to induce plasmid integration in comparison with that required in wild-type cells. This study suggests that NAD metabolism and intracellular NAM concentrations are important in Fob1-mediated rDNA recombination.
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, USA.
Kaushlendra
K
Tripathi
Nabil
N
Matmati
Shamsu
S
Zzaman
Caroline
C
Westwater
Bidyut K
BK
Mohanty
D015825:Chromosomes, Fungal / Q000235:genetics*; D004261:DNA Replication / Q000187:drug effects; D004271:DNA, Fungal / Q000187:drug effects; D004275:DNA, Ribosomal / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D016521:Electrophoresis, Gel, Pulsed-Field; D020868:Gene Silencing / Q000187:drug effects; D009536:Niacinamide / Q000494:pharmacology*; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic / Q000187:drug effects; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D056565:Sirtuin 2 / Q000235:genetics / Q000378:metabolism
D004271:DNA, Fungal; D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; C105579:FOB1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; D009536:Niacinamide; C087778:SIR2 protein, S cerevisiae; D056565:Sirtuin 2