Caloric restriction results in lifespan extension in yeast, worms, Drosophila and mammals. Yeast studies showed extension of lifespan in response to caloric restriction involves sir-2 the founding member of a widely conserved gene family encoding the sirtuins, enzymes with histone deacetylase activity. Like in yeast, Drosophila and mammals, over-expression of the C. elegans sirtuin gene sir-2.1 leads to extension of life span and deletion of the gene shortens life span (Tissenbaum & Guarente, 2001; Wang & Tissenbaum, 2006). Previously, a C-terminal translational gfp reporter fusion with sir-2.1 under the control of a 300 nucleotide intergenic promoter fragment (in strains HT809, HT810) was reported to reveal expression of sir-2.1 in most of the nerve cells in the head and the tail, and in the hypodermis (Wang & Tissenbaum, 2006). Recombineering allowed us to insert both a mCherry reporter gene just before the stop codon of sir-2.1 and a gfp reporter just before the stop codon of R11A8.6, within a 29.3kb fosmid genomic DNA clone. R11A8.6 is the first gene and sir-2.1 is the downstream gene in a two gene, 4.4kb operon. The mCherry expression pattern under normal growth conditions, with abundant food, indicated that sir-2.1 is indeed expressed, as previously described, in many nerves in the head. However, GFP was also seen, along with mCherry, in the gut and in head muscles. This reveals that expression of sir-2.1 is broader than previously described, fitting the expectation that the SIR2.1 function in influencing lifespan would be required in most cell types. Furthermore, the broader expression arises from transcription starting with the upstream gene of the sir-2.1 operon. SIR-2.1::mCherry is nuclear, consistent with the proposed function in controlling gene expression, while R11A8.6::GFP is cytoplasmic, consistent with the enzymatic role of the glutathione-S-transferase encoded by R11A8.6. Finally, and importantly, the expression of sir-2.1::mCherry and R11A8.6::gfp rises dramatically upon starvation. No such increase is seen with sir-2.1::gfp under the control of the intergenic promoter fragment suggesting that elevated sir-2.1 expression upon starvation depends on transcription across the operon from the upstream gene. Curiously, under starvation conditions, SIR-2.1::mCherry is not nuclear-localized, but is localized in puncta in the cytoplasm. Elevated transcription of sir-2.1 also corresponds with elevation of expression of sir-2.1 orthologues in other organisms upon starvation. We thank Heidi Tissenbaum for strains HT809, HT810. Tissenbaum & Guarente, Nature, 2001. 410, 227 Wang & Tissenbaum, Mech. Ageing Dev. 2006. 127, 48.

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