The Bayes significance was defined as 1- p(OR.1) when the posterior estimate of the OR was .1, and 1-p(OR,1) when the posterior estimate of the OR was ,1, and an association was deemed significant in the NECS, SICS, or the data aggregated from the two studies, if the Bayes significance was smaller than 0.05. This measure of significance is the posterior probability of the null hypothesis OR#1 (or OR$1) so that small values denotes strong evidence against the null hypothesis [32]. This analysis identified 18 significant SNPs (Table 3), that is more than twice the number expected by chance in 145 independent tests and two SNPs remained significant even after correcting the threshold for the number of tests. Furthermore, the probability that 18 SNPs could be simultaneously found significantly associated under the null hypothesis of no association is 0.0002, using the binomial distribution with n= 145, x= 18 and p= 0.05. An association that was significant in the aggregated NECS and SICS data was deemed replicated in either the AJCS or JCS studies if the same SNP was significant (Bayes significance ,0.05) with the same genetic model and consistent effects; or the same SNP did not reach statistical significance (Bayes significance $0.05), but the ORs in the different studies were in the same direction and when the data from the studies were aggregated, the association was significant. The rationale for the second condition is that both the AJCS and JC have smaller sample sizes, and therefore have less power compared to the NECS. However, consistent effects and increased significance when the aggregated data are analyzed show that the lack of association in the replication study is due to lack of power if effects are similar across different studies. This strategy has been used to increase the power of genetic association studies, see for example [51]. The results are in Tables 4, 5, 6 and 7.





We conducted a similar analysis stratified by gender but the limited sample sizes did not produce strongly significant results.

Linkage Disequilibrium (LD) Heatmaps

We used HaploView 4.1 to create the LD heatmaps and LD displays were generated using the D9 color scheme where white represents D9= 0, red represents D9= 1, and different shades of red represent 0,D9,1 (Figures 1 and 4).

Lifespan Measurements in C. elegans

To synchronize worms for lifespan, eggs were isolated (N2, adr- 1, adr-2, adr-1;adr-2, rde-1, rde-4, adr-1;adr-2;rde-1) and synchronized by hatching overnight in the absence of food at 20C. Synchronized L1 larvae were counted and plated (10 worms/plate, n =60) on Escherichia coli bacterial lawns (OP50) on NGM media and allowed to develop to L4-stage larvae at 20C. 5-fluorodeoxyur- idine (FudR) solution was added to a final concentration of 0.1mg/

OR for rs2838809 (Posteri ar density) OR for rs2838810 (Posterior density) OR for rs2838816 (Posterior density) 0.0 10 20 0.0 10 20 30 00 10 20OR=0.11 (0.39:1.26) OR=1.43 (0.82:2.67) OR=0.71 (0.39;1.26)P(OR>1)=0.12 P(OR<1)=0.11 P(OR>1)=0.12

Figure 3. Posterior densities of ORs. Posterior densities of the ORs for the 3 SNPs in ADARB1 with rare alleles and moderate effects in the data aggregated from NECS, SICS and JCS. Significant associations would results in posterior densities not overlapping 1 and definite evidence of either an OR,1 or an OR.1, while all of the three densities have heavy tails and do not provide definite evidence against the null hypothesis of no association. doi:10.1371/journal.pone.0008210.g003

ml to prevent reproduction. Worms were kept at 20C and monitored by counting on alternate days. Lifespan was defined the first day of adulthood (adult lifespan = 0) to death. Aside reduced lifespan, the worms appeared normal in size and

behavior, consistent with previous reports on adr mutant strains [36]. We did observe altered chemotaxis during routine passage of the worms to maintain stocks, as previously noted [36]. We also noted reduced progeny viability (data not shown). However, these

we ae Fu ebersesben IN BhOvEZzbS) 2ee zseleezss 2s BCete NFS zeigzozs!vercesiss < I xX 0so Ss olsesees:onsesscss 2 Ge ~es . evssoezs:erssoeze 2<=xX-2X8 . sessoezs:nH Stssoezs) . 8 = eBABA SK" esoueecss‘esgleeTs ¢ S 2 =sxssoezs) . -          Block 2 (17 Ko)  2   Blocks i)NECS 10195620AJCS

erssoszs)sessoezssesozeczs)==  [Ssii le= 3pialFO shorezensszsereczeszesozozssvercesisenssoezs)sessoszs)esozeccs)——=Ie                 Block 2 (17 ke)Blocks 71)3Btock 4 (1 kb)12Block 3 (6K)8 9 0) 47“56- =“4 1 38>39        Block 2 (17 Ko)A 3     2 Hoek 1 7b) 726 SICS1016   NECSelorezetss 2 2zsezeezes =zesgsozs) =57    orseeecsserssoeze)sessoezs) xesousezes °verses! -9zossoszsi °‘yszooiss 0 3‘ozreos01ss -Siorezzis)1210zse1eezs)"1   EB»542 4191056 220JCSAJCS

Figure 4. Pattern of LD among the SNP in ADARB2(chromosome 10) that are associated with exceptional longevity. The four plots display the pattern of LD captured by the SNPs associated with exceptional longevity in ADARB2 (chromosome 10) in the NECS, SICS, AJCS and JCS data. The intensity of red cells represents the strength of LD measured by D9. The LD pattern in the NECS, SICS and AJCS subjects are very similar but differ substantially from the pattern of LD in the JCS subjects in which two SNPs become almost monomorphic (rs884949 and rs2387653). Highlighted in red are the SNPs that replicate the results in the AJCS and JCS subjects. doi:10.1371/journal.pone.0008210.g004

rs17294019

183788157

P(A)0.80 082 0.84 0.86 0.88 0.90rs17294019100-105Age 106-119pia)0.640.601s3788157100-105 106-119Age