Association Analyses of Insulin Signaling Pathway Gene Polymorphisms with Healthy Aging and Longevity in Americans of Japanese Ancestry

Brian J. Morris, Timothy A. Donlon, Qimei He, John S. Grove, Kamal H. Masaki, Ayako Elliott, D. Craig Willcox, and Bradley J. Willcox

Honolulu Heart Program (HHP)/Honolulu-Asia Aging Study (HAAS), Kuakini Medical Center, Hawaii. Department of Geriatric Medicine, John A. Burns School of Medicine, University of Hawaii, Honolulu. Clinical Genomics Laboratory, School of Medical Sciences, University of Sydney, New South Wales. Public Health Sciences, John A. Burns School of Medicine, University of Hawaii at Manoa, Honolulu. Department of Human Welfare, Okinawa International University, Ginowan, Japan.

Address correspondence to Brian J. Morris, DSc PhD, Honolulu Heart Program (HHP)/Honolulu-Asia Aging Study (HAAS), Kuakini Medical Center, 347 North Kuakini Street, HPM-9, Honolulu, Hawaii 96817. Email: brian.morris@sydney.edu.au

Evidence from model organisms suggests that the insulin/IGF-1 signaling pathway has an important and evolutionarily conserved influence over the rate of aging and longevity. In humans, the FOXO3 gene is the only widely replicated insulin/IGF-1 signaling pathway gene associated with longevity across multiple populations. Therefore, we conducted a nested case-control study of other insulin/IGF-1 signaling genes and longevity using a large homogeneous long-lived population of American men of Japanese ancestry well-characterized for aging phenotypes. Genotyping was performed for single nucleotide polymorphisms tagging most of the genetic variation across several genes in the insulin/IGF-1 signaling pathway or related gene networks that may be influenced by FOXO3, namely, ATF4, CBL, CDKN2, EXO1, and JUN. Two initial marginal associations with longevity did not remain significant after correction for multiple comparisons, nor were they correlated with aging-related phenotypes.

Key Words: Longevity Molecular genetics Insulin signaling genes Human.

Received February 11, 2013; Accepted May 11, 2013

Heritability of extreme longevity is estimated as 0.33 and 0.48 in male and female centenarians (1). Although centenarians are enriched for longevity single nucleotide polymorphisms (SNPs) (2), few have been identified (3,4), possibly because of small effect sizes (5). 

The insulin/IGF-1 signaling pathway is an example of an evolutionarily conserved biological pathway that affects aging (6,7). In humans, a finding of association of genetic variation in the insulin/IGF-1 signaling pathway gene FOXO3A with extreme old age in our cohort of American men of Japanese ancestry (8) has now been consistently replicated in other populations for both sexes.

Methods

Here we report our findings from an exploratory study of SNPs in other evolutionarily conserved insulin/IGF-1 signaling pathway genes chosen on the basis of model organism data and human studies, namely, those encoding (i) activating transcription factor 4, which activates stress-resistance genes (9), transcription of the asparagine synthase gene (ASNS) in response to nutrient stress (10), and suppresses the cell senescence/lifespan determining components INK4A/ArF (locus CDKN2A) (11); (ii) Casitas B-cell lymphoma protein, which regulates whole-body energy expenditure and insulin action by suppression of the insulin receptor system. Study Population

The study used a nested case-control design and participants drawn from the HHP/HAAS population described previously (8). These were American men of Japanese ancestry recruited in Oahu in 1965-1968 at ages 45-68 years and followed with regular examinations and blood work until the present, or death. Cases (longevity phenotype) were defined as HHP/HAAS participants who had survived to beyond the upper 1% of the 1910 U.S. birth cohort-specific survival (minimum 95 years of age [19]; n = 213 as of August 2007, 176 died [age of death = 97.5 ± 2.1 SD; range 95-106 years] and 37 were still alive [mean age = 98.7 ± 2.1 SD; range 97-106 years]). Controls consisted of 402 individuals from the HHP/HAAS cohort who had died near the mean for the 1910 U.S. birth cohort-specific survival for middle-aged men (77 years). To achieve a case:control ratio of approximately 1:2, we sampled the HHP/HAAS study populationNucleotide Total Number of Minor Allele Frequency of Cases/ Polymorphism ID Alleles Case/Controls Controls; n (%; 95% CI) 2 p-Value 

ATF4 rs1048310 G/T 208/399 44/82 (10.6/10.3; 0.03 to 0.04) 0.027 0.87 
rs6519194 A/G 206/397 12/17 (2.9/2.1; 0.01 to 0.029) 0.69 0.41 
rs738288 A/G 205/396 4/11 (0.98/1.4; 0.02 to 0.011) 0.37 0.54 
CBL rs6589722 A/C 209/402 80/111 (19/14; 0.009 to 0.099) 5.9 0.015 
rs12786104 G/T 210/398 181/380 (43/47; 0.10 to 0.012) 2.4 0.12 
rs11217184 A/G 212/396 157/299 (37/38; 0.06 to 0.05) 0.06 0.80 
rs2249466 C/T 207/394 136/256 (33/32; 0.05 to 0.06) 0.02 0.90 
rs7914 A/G 190/376 150/306 (39/41; 0.07 to 0.048) 0.16 0.69 
rs2511837 A/G 212/398 143/236 (34/30; <sup>1</sup>/sub>0.01 to 0.096) 2.1 0.14 
CDKN2B rs10811661 C/T 213/402 182/380 (43/47; 0.10 to 0.013) 2.3 0.13 
EXO1 rs1776180 C/G 206/401 11/18 (5.3/4.4; 0.01 to 0.025) 3.6 0.16 
JUN rs2999131 C/G 208/391 132/226 (32/29; 0.03 to 0.083) 1.0 0.31 
rs1815274 C/T 212/401 164/279 (37/35; <sup>2</sup>/sub>0.02 to 0.096) 1.8 0.18 
rs17356414 T/C 212/401 113/252 (27/20; <sup>3</sup>/sub>0.010 to 0.006) 3.0 0.082 
rs1853432 A/T 213/398 184/359 (43/45; <sup>4</sup>/sub>0.08 to 0.039) 0.41 0.52 

Note: CI = confidence interval.

One-way ANOVA did not detect any relationship between factors associated with insulin sensitivity, a potential intermediate phenotype of longevity, and genotype. For non-normally distributed variables, we used log transformation to approximate a normal distribution.