Figure 6. Glycogen accumulation is responsible for glucose-mediated life shortening.

(a) Worms lacking glycogen deposition live much longer on a high glucose diet. N2 worms were fed on NGM plates seeded with E. coli HT115 harbouring either empty vector (black and red curves) or plasmids expressing either gsy-1 (blue and green curves) or pyg-1 (cyan and magenta curves) RNAi. L4 stage animals were transferred to NGM plates without (black, blue and cyan curves) or with (red, green and magenta curves) glucose. The graph shows the average of three independent experiments (see also Supplementary Table 2). (b) Glycogen content in gsy-1 and pyg-1 knockdown worms. Animals were grown on NGM (green bars), NGM+2% glucose (red bars) or NGM+2% glucose+5 mM diamide (orange bars) agar plates as in panel a or c. Seven-day-old worms were then picked, washed and the amount of glycogen determined and normalized to the total protein. The graph shows the average±s.d. of at least three independent experiments (see also Supplementary Table 3). (c) Diamide fails to extend the short lifespan of glycogen phosphorylase-deficient worms fed on high glucose diet. N2 worms were fed on NGM plates seeded with E. coli HT115 harbouring either empty vector (black curve) or plasmid vector expressing either gsy-1 (red curve) or pyg-1 (green curve) RNAi. L4 stage animals were transferred to NGM plates with 2% glucose and 5 mM diamide. The graph shows the average of three independent experiments (see also Supplementary Table 2). (d) Diamide does not efficiently deplete glycogen in pyg-1-deficient worms. A representative image of four-day-old worms stained for glycogen with iodine is shown. Scale bar 0.15 mm. Experimental conditions were as in panel c.