TCER-1 Links Reproductive Signals to Longevity Figure 5. tcer-1 overexpression causes increased transcription of DAF-16/FOXO-target genes. (A) mRNA levels of DAF-16/FOXO-targets compared between tcer-1(2) (green bars) and tcer-1(2); tcer-1 OE (red bars) animals. Day 1 adults grown under similar conditions were analyzed by Q-PCR. Data shown here is combined from six independent biological replicates, each comprising 2–4 technical repeats. The genes tested are displayed on the X-axis and relative expression levels are on the Y-axis. Error bars display standard error of the mean. The asterisks represent a change in expression between the two strains with an unpaired, two-tailed t-test with a P value,0.05 (*) or ,0.005 (**). (B) Lifespan extension mediated by tcer-1 overexpression depends on daf-16. Wild type (N2): m = 18.460.3, n = 66/98; daf-16(mu86): m = 17.560.2, n = 65/81, P vs. N2 = 0.03; tcer-1 OE: m = 21.160.2, n = 79/102, P vs. N2,0.0001; daf-16(mu86); tcer-1 OE: m = 17.660.2, n = 89/104, P vs. N2 = 0.54, P vs. daf-16(mu86) = 0.15, P vs. tcer-1 OE,0.0001. See Table S4 for multiple repeats of this experiment. doi:10.1371/journal.pgen.1000639.g005 tion in germline-defective animals, affected TCER-1 up-regulation (data not shown). However, in kri-1(ok1251) mutants, nuclear TCER-1::GFP levels were severely diminished in the intestine and were not increased following germ-cell ablation (Figure 7B–7F). kri-1(ok1251) did not alter TCER-1::GFP levels in any other tissues (Figure 7E and 7F), consistent with KRI-1’s presence only in the gastrointestinal tract [13]. Thus, DAF-16/FOXO nuclear local- ization and TCER-1 up-regulation share a requirement for KRI-1 in their response to germ-cell loss. As expected, kri-1(ok1251) also abolished the lifespan extension evoked by tcer-1 overexpression (Figure 7G; Table S4). KRI-1 is not required for DAF-16/FOXO nuclear localization or lifespan extension in daf-2 mutants [13]. Thus, KRI-1’s involvement in TCER-1 regulation explains, at least in part, why TCER-1 function is specific for the reproductive system. Discussion The finding that signals from reproductive tissues can influence the lifespan of an animal is fascinating and important because both reproduction and aging are such fundamental aspects of an animal’s life history. In this study, we have shown that a predicted transcription elongation factor, TCER-1, is a key regulatory target of lifespan-extending signals from the repro- ductive system in C. elegans. TCER-1’s level in the animal is sensitive to cues from the reproductive system, and when overexpressed, TCER-1 can bypass the normal requirement for germline loss and extend the lifespan of fertile animals with an intact reproductive system. Our data indicate that TCER-1 extends lifespan by collaborat- ing with the DAF-16/FOXO transcription factor, which is also required for the loss of germ cells to extend lifespan [1]. DAF-16/ FOXO has an evolutionarily-conserved role in lifespan regulation [18,44], and is activated in many other circumstances that increase lifespan, including reduced insulin/IGF-1 signaling [8,11] or overexpression of the sirtuin SIR-2.1 [45], heat-shock factor HSF- 1 [46], AMP kinase [47] or c-Jun N-terminal kinase [48]. How such a core, conserved longevity factor is activated by many disparate stimuli is a central question in the biology of aging. In this study, we have shown that TCER-1 plays a key role in establishing a distinct pattern of DAF-16/FOXO-dependent gene expression specifically in response to signals from the germline. PLoS Genetics | www.plosgenetics.org 7 September 2009 | Volume 5 | Issue 9 | e1000639