Aging research has developed rapidly over the past decade, identifying individual genes and molecular mechanisms of the aging process through the use of model organisms and high throughput technologies. Calorie restriction (CR) is the most widely researched environmental manipulation that extends lifespan. Activation of the NAD+-dependent protein deacetylase Sir2 (Silent Information Regulator 2) has been proposed to mediate the beneficial effects of CR in the budding yeast Saccharomyces cerevisiae, as well as other organisms. Here, we show that in contrast to previous reports, Sir2 is not stimulated by CR to strengthen silencing of multiple reporter genes in the rDNA of S. cerevisiae. CR does modestly reduce the frequency of rDNA recombination, although in a SIR2-independent manner. CR-mediated repression of rDNA recombination also does not correlate with the silencing of Pol II-transcribed noncoding RNAs derived from the rDNA intergenic spacer, suggesting that additional silencing-independent pathways function in lifespan regulation. © 2009 The Authors Journal compilation © Blackwell Publishing Ltd/Anatomical Society of Great Britain and Ireland 2009.