Sphingolipids Control Lifespan Figure 2. Down-regulating sphingolipid synthesis increases CLS. (A) Growth rate of tetO7-LCB1 cells (RCD956) treated with Dox to down- regulate expression of the tetO7 promoter and reduce the rate of sphingolipid synthesis. (B) Growth rate of tetO7-LCB2 cells (RCD957) +/2 Dox treatment. (C) CLS of tetO7-LCB1 cells +/2 Dox treatment. Data show the mean 6 SEM of surviving cells (* p,0.05, ** p,0.01, No Dox vs 100 ng/ml Dox). (D) CLS of tetO7-LCB2 cells +/2 Dox treatment. Assays are similar to those presented in (C). (E) Verification that Dox-treatment extends CLS by reducing expression of the tetO7-LCB1 target gene. CLS of tetO7-LCB1 cells transformed with an empty vector (Vec1 = pRS315ADE2) or a vector carrying LCB1 (pLCB1 = pRS315ADE2-LCB1-5) is shown. Data represent the mean 6 SEM of surviving cells (* p,0.05, ** p,0.01, for Vec1, 100 ng/ml PLoS Genetics | www.plosgenetics.org 4 February 2012 | Volume 8 | Issue 2 | e1002493