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Originally published as Genetics Published Articles Ahead of Print on September 9, 2008.

Genetics, Vol. 180, 797-810, October 2008, Copyright © 2008
doi:10.1534/genetics.108.091090

Pnc1p-Mediated Nicotinamide Clearance Modifies the Epigenetic Properties of rDNA Silencing in Saccharomyces cerevisiae

Julie M. McClure1, Christopher M. Gallo1, Daniel L. Smith, Jr., Mirela Matecic, Robert D. Hontz, Stephen W. Buck, Frances G. Racette and Jeffrey S. Smith2

Department of Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia 22908

2 Corresponding author: Department of Biochemistry and Molecular Genetics, University of Virginia Health System, 1340 Jefferson Park Ave., Jordan Hall, Box 800733, Charlottesville, VA 22908.
E-mail: jss5y{at}virginia.edu

The histone deacetylase activity of Sir2p is dependent on NAD+ and inhibited by nicotinamide (NAM). As a result, Sir2p-regulated processes in Saccharomyces cerevisiae such as silencing and replicative aging are susceptible to alterations in cellular NAD+ and NAM levels. We have determined that high concentrations of NAM in the growth medium elevate the intracellular NAD+ concentration through a mechanism that is partially dependent on NPT1, an important gene in the Preiss–Handler NAD+ salvage pathway. Overexpression of the nicotinamidase, Pnc1p, prevents inhibition of Sir2p by the excess NAM while maintaining the elevated NAD+ concentration. This growth condition alters the epigenetics of rDNA silencing, such that repression of a URA3 reporter gene located at the rDNA induces growth on media that either lacks uracil or contains 5-fluoroorotic acid (5-FOA), an unusual dual phenotype that is reminiscent of telomeric silencing (TPE) of URA3. Despite the similarities to TPE, the modified rDNA silencing phenotype does not require the SIR complex. Instead, it retains key characteristics of typical rDNA silencing, including RENT and Pol I dependence, as well as a requirement for the Preiss–Handler NAD+ salvage pathway. Exogenous nicotinamide can therefore have negative or positive impacts on rDNA silencing, depending on the PNC1 expression level.




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