We have performed the largest genome-wide association meta-analysis for human longevity, in which a novel locus on chromosome 5q33.3 associating with survival beyond 90 years was identified.

The minor allele of rs2149954 (T) promotes human longevity by reducing the risk of mortality owing to stroke and non- cardiovascular causes. In addition, this allele has previously been associated with low blood pressure, which may explain the protection from CVD mortality risk in middle age. At ages above 80years,however,low SBPassociateswithincreased mor- tality (37,38). Hence, the observed blood pressure-independent ≥ association of the minor allele with mortality 75 years may be due to pleiotropic effects on other mortality-related clinical para- meters. Examination of publically available data of several large GWAS consortia for association of the locus with parameters related to glucose and fat metabolism provided as yet no clues for other potentially involved mechanisms.

Phenotypic characterization and pathway analysis

Inanattempttoidentifytheunderlyingmechanismbywhichthis novel longevity locus at chromosome 5q33.3 could influence human longevity, we examined rs2149954 in the published data of several large GWAS consortia for association with meta- bolic traits in generally middle-aged individuals. None of the investigated traits, i.e. 2 h glucose (OGTT), Hb Ac, fasting 1 glucose, fasting insulin, insulin resistance (HOMA-IR), b-cell activity (HOMA-B), total/HDL/LDL cholesterol, triglycerides and type 2 diabetes (26–32), demonstrated evidence for associ- ation (all P . 0.05) with rs2149954 (Supplementary Material, Tables S8 and S9).

Table 2. Results of the discovery phase, replication phase and joint analysis of cases aged ≥90 years

Locus Lead SNP Chromosome Position Candidate/closest gene EA Analysis n EAF OR 95% CI P I2 (%) P het Cases Controls Cases Controls 1q43 rs1625040 1 235 213 002 MTR, RYR2 A Discovery 5406 15 112 0.176 0.150 1.18 1.10–1.26 6.53 × 1026 Replication 7310 60 914 0.175 0.182 1.05 0.99–1.10 0.065 Joint 12 716 76 026 1.10 1.05–1.14 2.60 × 1025 9.3 0.343 4q22.2 rs4693331 4 94 760 609 GRID2 C Discovery 5406 15 112 0.416 0.444 0.89 0.84–0.93 6.63 × 1026 Replication 7267 60 324 0.449 0.440 1.03 0.99–1.07 0.095 Joint 12 673 75 436 0.97 0.94–1.00 0.139 61.3 3.51 × 1024 4q27 rs13114426 4 120 942 533 PDE5A, MAD2L1 T Discovery 5406 15 112 0.381 0.405 0.88 0.84–0.92 2.11 × 1026 Replication 7305 60 932 0.369 0.351 0.98 0.94–1.02 0.336 Joint 12 711 76 044 0.94 0.91–0.97 1.95 × 1024 32.5 0.090 5q33.3 rs2149954 5 157 753 180 EBF1 T Discovery 5406 15 112 0.396 0.360 1.14 1.09–1.21 1.85 × 1026 Replication 7298 60 262 0.374 0.352 1.07 1.03–1.12 5.98 × 1024 Joint 12 704 75 374 1.10 1.06–1.14 1.74 × 1028 28.5 0.125 7p14.2 rs11977641 7 36 761 949 AOAH, ELMO1 C Discovery 5406 15 112 0.062 0.076 0.78 0.70–0.87 7.31 × 1026 Replication 3049 4805 0.071 0.073 0.93 0.82–1.06 0.226 Joint 8455 19 917 0.84 0.77–0.91 1.57 × 1025 50.2 0.010 10q23.33 rs4466755 10 96 622 243 CYP2C19, CYP2C9 T Discovery 5406 15 112 0.455 0.445 1.13 1.07–1.18 1.30 × 1025 Replication 7326 61 105 0.477 0.508 0.98 0.94–1.02 0.208 Joint 12 732 76 217 1.03 1.00–1.07 0.087 55.4 0.002 12q15 rs11834614 12 67 197 344 MDM1, RAP1B C Discovery 5406 15 112 0.138 0.155 0.85 0.79–0.91 9.94 × 1026 Replication 7272 60 210 0.165 0.173 1.01 0.96–1.07 0.603 Joint 12 678 75 322 0.95 0.91–0.99 0.023 43.9 0.024 14q23.2 rs2784505 14 61 501 766 SYT16 G Discovery 5406 15 112 0.080 0.067 1.23 1.11–1.35 8.87 × 1025 Replication 7323 60 979 0.070 0.066 1.10 1.02–1.19 0.012 Joint 12 729 76 091 1.15 1.08–1.22 9.47 × 1026 28.3 0.127 17p13.1 rs940850 17 8 870 805 NTN1 T Discovery 5405 15 112 0.072 0.093 0.78 0.70–0.87 4.93 × 1026 Replication 7276 60 146 0.109 0.118 1.03 0.97–1.10 0.318 Joint 12 681 75 258 0.95 0.90–1.01 0.111 63.7 1.32 × 1024 17q23.2 rs2109265 17 58 307 001 MARCH10, TANC2 A Discovery 5406 15 112 0.443 0.420 1.13 1.08–1.19 3.34 × 1026 Replication 7307 60 672 0.453 0.465 1.01 0.97–1.05 0.671 Joint 12 713 75 784 1.06 1.02–1.09 0.001 34.7 0.074 19q13.32 rs4420638a 19 50 114 786 APOE G Discovery 5405 15 102 0.145 0.195 0.64 0.59–0.70 4.09 × 10221 Replication 4861 57 126 0.165 0.202 0.77 0.72–0.82 2.95 × 10218 Joint 10 266 72 228 0.72 0.68–0.76 3.40 × 10236 70.1 3.69 × 1025 20q13.2 rs8126377 20 51 590 254 TSHZ2, ZNF217 G Discovery 5209 14 893 0.057 0.068 0.75 0.66–0.85 3.38 × 1025 Replication 7278 60 647 0.063 0.054 1.04 0.95–1.13 0.309 Joint 12 487 75 540 0.94 0.87–1.00 0.117 58.1 0.001



EA, effect allele; EAF, effect allele frequency after pooling the data of all analyzed individuals; OR, odds ratio for the effect allele; 95% CI, 95% confidence heterogeneity. aGenotyping of this SNP with the Sequenom MassARRAY system for the replication phase was unsuccessful. The SNPs in bold overlap with Table 1.

—logio(p-value)     Position on chr19 (Mb)  we ROT eaePie cup ~anpase cLocyso= — ~ ony}Tome srpgig-= ~mazpenaarogt= cer ~exgeazswocyer-=aropeesroge50.2 50.4(QW/W9) aye UoneUIquosey—logio(p-value)    157.6 157.8Position on chr5 (Mb) (w/w) e721 uopeuiquiossy

Figure3.Regionalassociationplotsforthechromosome19q13.32and5q33.3loci.Resultsofthediscovery-phaseanalysisofchromosome19q13.32(A)and5q33.3 (B) in cases aged ≥90 years, generated using LocusZoom (http://csg.sph.umich.edu/locuszoom/) (22). For the two SNPs taken forward to the replication phase (rs4420638 and rs2149954), the results of the joint analysis are plotted. The color of the SNPs is based on the LD with the lead SNP (shown in purple). The blue peaksrepresenttherecombinationratesbasedonHapMapPhaseI+IICEUrelease22(hg18/build36),andtheRefSeqgenesintheregionareshowninthelowerpanel.