Fig. 4.

Characterization of SIR2 function in Candida aging. (A) Lower glucose concentration lengthens the lifespan of C. albicans. Left panel, strain SC5314; right panel, strain BWP17. Mean lifespan and sample size of SC5314: 2% glucose 19.1 (n = 54), 0.1% glucose 23.7 (n = 60); BWP17: 2% glucose 28.0 (n = 50), 0.1% glucose 36.8 (n = 50). (B) Left panel, deletion of one copy of SGS1 causes a shortened lifespan. Mean lifespan and sample size are: BWP17 (wild-type) 28.1 (n = 72), sgs1Δ/SGS1 12.6 (n = 67). Middle panel, reduced dosage of Sir2 decreases lifespan. Mean lifespan and sample size are: BWP17 (wild-type) 27.4 (n = 60), sir2Δ/SIR2 18.2 (n = 60), sir2Δ/sir2Δ 11.9 (n = 60). Right panel, insertion of an extra copy of SIR2 results in lifespan extension. Mean lifespan and sample size are: BWP17 (vector) 22.4 (n = 60), SIR2/ADE2 27.8 (n = 60). (C) Haplo-insufficiency of SIR2 causes a reduction of rDNA silence. Cells from logarithmic culture were 10-fold serially diluted, spotted onto plates, incubated at 30 °C for 2 days and photographed. The isogenic strains were labeled on the left. Left panel, cells grown on YC plate; right panel, cells grown on YC+5-FOA plate. BWP17 UR represents the strain with a URA3 gene inserted in rDNA loci. (D) Detection of extra-chromosomal rDNA molecules using pulsed field gel electrophoresis. Left panel, ethidium bromide staining of the PFGE gel; middle panel, Southern blot examination with an rDNA probe. The isogenic strains were labeled on the top and each lane represents an independent clone of the corresponding strain. Marker, Low Range PFG marker (New England BioLabs); Saccharomyces cerevisiae (strain BY4742) chromosomes are used as another marker. Arrows indicate the extra-chromosomal rDNA molecules. Right panel shows the quantification of extra-chromosomal rDNA molecules (ERMs). The relative amount of ERMs in BWP17 strain was assigned a value of 1. Error bars represent standard deviations of two independent clones.