Association of the FOXO3A Locus with Extreme Longevity in a Southern Italian Centenarian Study

Chiara Viviani Anselmi,1* Alberto Malovini,1,2* Roberta Roncarati,1 Valeria Novelli,1 Francesco Villa,1 Gianluigi Condorelli,1 Riccardo Bellazzi,2 and Annibale Alessandro Puca,1

Abstract

A number of potential candidate genes in a variety of biological pathways have been associated with longevity in model organisms. Many of these genes have human homologs and thus have the potential to provide in- sights into human longevity. Recently, several studies suggested that FOXO3A functions as a key bridge for various signaling pathways that influence aging and longevity. Interestingly, Willcox and colleagues identified several variants that displayed significant genotype–gender interaction in male human longevity. In particu- lar, a nested case–control study was performed in an ethnic Japanese population in Hawaii, and five candidate longevity genes were chosen based on links to the insulin–insulin-like growth factor-1 (IGF-1) signaling path- way. In the Willcox study, the investigated genetic variations (rs2802292, rs2764264, and rs13217795) within the FOXO3A gene were significantly associated with longevity in male centenarians. We validated the association of FOXO3A polymorphisms with extreme longevity in males from the Southern Italian Centenarian Study. Par-   ticularly, rs2802288, a proxy of rs2802292, showed the best allelic association—minor allele frequency (MAF)   7a 0.49; p 0.003; odds ratio (OR) 1.51; 95% confidence interval (CI), 1.15–1.98). Furthermore, we undertook a meta-analysis to explore the significance of rs2802292 association with longevity by combining the association results of the current study and the findings coming from the Willcox et al. investigation. Our data point to a key role of FOXO3A in human longevity and confirm the feasibility of the identification of such genes with centenarian–controls studies. Moreover, we hypothesize the susceptibility to the longevity phenotype may well be the result of complex interactions involving genes and environmental factors but also gender.

Introduction

for several age-related diseases that cause significant mor- bidity and mortality among older people.

GING IS A MULTIFACTORIAL AND COMPLEX by stochastic interactions (random damage to vital mol- ecules), extrinsic interventions (such as diet and caloric re- striction), and intrinsic/genetic alterations. A number of cen- tenarian (people aged 100 years old or older) studies exploit the strong selection of favorable genotypes in exceptionally aged individuals to study candidate genes and to perform genome wide analyses. Replications of observed associations of genotypes with longevity, coupled with functional stud- ies to define mechanisms whereby specific genotypes influ- ence lifespan-associated phenotypes, are essential to delin- eate true positive genetic findings. Such findings could potentially lead to preventive and therapeutic interventions

Numerous genes have been identified that are either pos- itively or negatively selected in the centenarian population as a consequence of a demographic selection,1–3 but no con- sistent replications have been observed in independent pop- ulation, with the exception of Apolipoprotein E (APOE).4,5 In addition, animal studies have provided insights into the types of genes that can be involved in the regulation of life- span. The first longevity mutant to be identified was the Caenorhabditis elegans gene PAX2 age-1.6 This mutant encodes phosphatidylinositol-3-kinase (PI3K),7 which has a key role in a signaling pathway that is homologous to the mammalian insulin–insulin-like growth factor-1 (IGF-1) pathway. Ge-

1Unit of Genetics and Cardiovascular Research Institute, Istituto Ricovero Cura Carattere Scientifico “Multimedica”, Sesto S. Giovanni,

2 Department of Computer Engineering and Systems Science, University of Pavia, Pavia, Italy. *These authors contributed equally to this work.

netic studies in C. elegans have demonstrated that activation of the PIK3/Akt pathway by IGF-1 suppresses activity of the DAF-16 forkhead transcription factor, the nematode ortholog of mammalian FOXO proteins. FOXO transcription factors belong to the Forkhead family of proteins, a family charac- terized by a conserved DNA-binding domain (Forkhead box) for phosphorylation by the survival kinase Akt.8–10 Particu- larly, in mammals the FOXO subgroup (FOXO1, FOXO3, FOXO4, and FOXO6) promotes the expression of numerous downstream genes that mediate stress resistance, innate im- munity, metabolic processes, and toxin degradation.11–15

rs13217795 (p   0.0006), and rs2802292 (p r<( 0.0001).22 The aim of our study was to support the potential role of FOXO3A as a key component that influences longevity with a case–control study. We screened 480 long-living “cases” (281 males and 199 females) and 335 young controls (195 males, 140 females) from an isolated and homogeneous pop- ulation of southern Italy (Southern Italian Centenarian Study, SICS) with an Illumina BeadChip 300K system, and we focused on the FOXO3A gene. As previously described, we hypothesized that gender might influence the association between FOXO3A and longevity; thus, we analyzed males and females separately. We applied a powerful method for correcting for stratification of our population. Last, we ex- ploited a meta-analysis approach to pool Willcox’s findings to find evidence for a statistical association for the SNP rs2802292.